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94-400: See text Potyvirus is a genus of positive-strand RNA viruses (named after its type species, Potato virus Y (PVY) ) in the family Potyviridae . Plants serve as natural hosts. Like begomoviruses , members of this genus may cause significant losses in agricultural, pastoral, horticultural , and ornamental crops. More than 200 species of aphids spread potyviruses, and most are from

188-455: A gem-diol intermediate. Carboxylation and hydration have been proposed as either a single concerted step or as two sequential steps. Concerted mechanism is supported by the proximity of the water molecule to C3 of RuBP in multiple crystal structures. Within the spinach structure, other residues are well placed to aid in the hydration step as they are within hydrogen bonding distance of the water molecule. The gem-diol intermediate cleaves at

282-438: A genome and as messenger RNA ; it can be directly translated into protein in the host cell by host ribosomes . The first proteins to be expressed after infection serve genome replication functions; they recruit the positive-strand viral genome to viral replication complexes formed in association with intracellular membranes. These complexes contain proteins of both viral and host cell origin, and may be associated with

376-501: A lysine in the active site (forming a carbamate ). Mg operates by driving deprotonation of the Lys210 residue, causing the Lys residue to rotate by 120 degrees to the trans conformer, decreasing the distance between the nitrogen of Lys and the carbon of CO 2 . The close proximity allows for the formation of a covalent bond, resulting in the carbamate. Mg is first enabled to bind to

470-462: A transmembrane domain , 6K1 is thought to contribute to virus-induced vesicle formation. 6K2 (~6 kDa) is a transmembrane protein which rearranges host membranes into virus-induced membrane structures. It interacts with various ER exit site proteins to produce vesicular and tubular extensions which eventually mature into replication vesicles. 6K2 has three main domains: the N-terminal domain which

564-425: A central hollow cylinder from which laminate sheets radiate outward and fold in on themselves in a pattern often described as " pinwheels ". These inclusions are easily seen in transmission electron micrographs of infected tissues and were historically used as a diagnostic criterion for potyvirus infections. CI inclusions are a major component of the potyviral movement complex which is assembled at plasmodesmata . CI

658-494: A compromise between specificity and reaction rate. It has been also suggested that the oxygenase reaction of RuBisCO prevents CO 2 depletion near its active sites and provides the maintenance of the chloroplast redox state. Since photosynthesis is the single most effective natural regulator of carbon dioxide in the Earth's atmosphere , a biochemical model of RuBisCO reaction is used as the core module of climate change models. Thus,

752-403: A correct model of this reaction is essential to the basic understanding of the relations and interactions of environmental models. There currently are very few effective methods for expressing functional plant Rubisco in bacterial hosts for genetic manipulation studies. This is largely due to Rubisco's requirement of complex cellular machinery for its biogenesis and metabolic maintenance including

846-488: A diversity of plant lineages, ancestral C 3 -type RuBisCO evolved to have faster turnover of CO 2 in exchange for lower specificity as a result of the greater localization of CO 2 from the mesophyll cells into the bundle sheath cells . This was achieved through enhancement of conformational flexibility of the “open-closed” transition in the Calvin cycle . Laboratory-based phylogenetic studies have shown that this evolution

940-645: A high level of homology with the picornaviral 3C protease . VPg (~22 kDa) is covalently attched to the 5' end of the viral genomic RNA through uridylation and is thought to act as a primer for viral genome replication similarly to the VPg proteins of picornaviridae . It is a highly disordered protein and its flexibility has been suggested to allow it to interact with many other viral proteins. VPg also interacts with various host proteins including eukaryotic initiation factor 4E (eIF4E), eukaryotic elongation factor 1A (eEF1A), and poly(A)-binding protein (PABP). NIb (~59 kDa)

1034-406: A high optimal pH (can be >9.0, depending on the magnesium ion concentration) and, thus, becomes "activated" by the introduction of carbon dioxide and magnesium to the active sites as described above. In plants and some algae, another enzyme, RuBisCO activase (Rca, GO:0046863 , P10896 ), is required to allow the rapid formation of the critical carbamate in the active site of RuBisCO. This

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1128-418: A higher affinity for CO 2 . The process first makes a 4-carbon intermediate compound, hence the name C 4 plants, which is shuttled into a site of C 3 photosynthesis then decarboxylated, releasing CO 2 to boost the concentration of CO 2 . Crassulacean acid metabolism (CAM) plants keep their stomata closed during the day, which conserves water but prevents the light-independent reactions (a.k.a.

1222-553: A sister clade in relation to Lenarviricota . The third phylum that contains +ssRNA viruses is Pisuviricota , which has been informally called the "picornavirus supergroup". The phylum contains a large assemblage of eukaryotic viruses known to infect animals, plants, fungi, and protists. The phylum contains three classes, two of which contain only +ssRNA viruses: Pisoniviricetes , which contains nidoviruses , picornaviruses , and sobeliviruses , and Stelpaviricetes , which contains potyviruses and astroviruses . The third class

1316-447: A strategy to increase crop yields. Approaches under investigation include transferring RuBisCO genes from one organism into another organism, engineering Rubisco activase from thermophilic cyanobacteria into temperature sensitive plants, increasing the level of expression of RuBisCO subunits, expressing RuBisCO small chains from the chloroplast DNA , and altering RuBisCO genes to increase specificity for carbon dioxide or otherwise increase

1410-730: A variety of host organelles and the sources differ between potyvirus species. Some membrane sources include the ER , chloroplasts , Golgi apparatus , and vacuoles . The exact replication mechanism is not known but it involves a negative sense RNA intermediate and requires both viral and host proteins. Viral proteins detected in replication complexes include HC-Pro, P3, 6K1, 6K2, CI, VPg, NIa-Pro, and NIb. Host factors present in replication vesicles include eIF4A and several heat shock proteins . Like most plant viruses, potyviruses have evolved to move from one plant cell to another through plasmodesmata . However, unlike some well-studied plant viruses, such as

1504-399: Is Duplopiviricetes , whose members are double-stranded RNA viruses that are descended from +ssRNA viruses. Ribulose-1,5-bisphosphate carboxylase RuBisCO is important biologically because it catalyzes the primary chemical reaction by which inorganic carbon enters the biosphere . While many autotrophic bacteria and archaea fix carbon via the reductive acetyl CoA pathway ,

1598-580: Is a cysteine protease which cleaves a glycine -glycine dipeptide at its own C-terminus . It also interacts with eukaryotic initiation factor 4 (eIF4). It acts as a viral RNA silencing suppressor through its interactions with host AGO proteins. HC-Pro's activity is regulated by the adjacent P1 protein: before P1 cleaves itself off the P1-HC-Pro intermediate, the P1 terminus reduces HC-Pro's RNA silencing suppression activity. The rate of P1 cleavage therefore regulates

1692-428: Is a serine protease which facilitates its own cleavage from the polyprotein at the P1-HC-Pro junction. P1 consists of a conserved C-terminal protease domain and an N-terminal region which has a high level of variation in sequence and length between potyvirus species but exhibits conserved patterns of intrinsic disorder . P1 is also promotes viral RNA replication, though it is not required for it. HC-Pro (~52 KDa)

1786-412: Is a superfamily II RNA-dependent RNA polymerase (RdRp) which polymerises viral RNA during replication. Like NIa, NIb forms inclusions in the host nucleus where it is transported due to its two nuclear localisation sequences . NIb has the three-domain "palm, thumb, and fingers" structure typical of RdRps. 6K1 (~6 kDa) the function is not known, but because it accumulates in replication vesicles and has

1880-497: Is about 1.15 × 10 nucleotide substitutions/site/year. Agriculture was introduced into Australia in the 18th century. This introduction also included plant pathogens. Thirty eight potyvirus species have been isolated in Australia. Eighteen potyviruses have been found only in Australia and are presumed to be endemic there. The remaining twenty appear to have been introduced with agriculture. Historically, potyvirus diagnostics relied on

1974-424: Is also required for viral replication and is present on replication membranes. Its exact contributions to replication are not clear but, as an RNA helicase, CI is likely facilitating replication by dismantling the secondary structures of viral RNA. NIa (~50 kDa) forms crystalline inclusions in the host nucleus. It is cleaved into NIa-Pro and VPg. NIa-Pro (~27 kDa) is a cysteine protease which processes most of

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2068-477: Is encoded by the chloroplast DNA in plants. There are typically several related small-chain genes in the nucleus of plant cells, and the small chains are imported to the stromal compartment of chloroplasts from the cytosol by crossing the outer chloroplast membrane . The enzymatically active substrate ( ribulose 1,5-bisphosphate) binding sites are located in the large chains that form dimers in which amino acids from each large chain contribute to

2162-431: Is favored by an alkaline pH . The pH and the concentration of magnesium ions in the fluid compartment (in plants, the stroma of the chloroplast ) increases in the light. The role of changing pH and magnesium ion levels in the regulation of RuBisCO enzyme activity is discussed below . Once the carbamate is formed, His335 finalizes the activation by returning to its initial position through thermal fluctuation. RuBisCO

2256-473: Is further disrupted by viral proteases degrading components required to initiate translation of cellular mRNA. All positive-strand RNA virus genomes encode RNA-dependent RNA polymerase , a viral protein that synthesizes RNA from an RNA template. Host cell proteins recruited by +ssRNA viruses during replication include RNA-binding proteins , chaperone proteins , and membrane remodeling and lipid synthesis proteins, which collectively participate in exploiting

2350-432: Is inhibited (or protected from hydrolysis) by a competitive inhibitor synthesized by these plants, a substrate analog 2-carboxy-D-arabitinol 1-phosphate (CA1P). CA1P binds tightly to the active site of carbamylated RuBisCO and inhibits catalytic activity to an even greater extent. CA1P has also been shown to keep RuBisCO in a conformation that is protected from proteolysis . In the light, RuBisCO activase also promotes

2444-678: Is not known whether cap-independent translation is an important translation mechanism for potyviruses. Like many other positive strand RNA viruses , potyvirus replication is heavily associated with host membranes. The viral 6K2 protein coordinates the rearrangement of host membranes into various infection-associated structures which, depending on the potyvirus species, can include anything from small round viral vesicles to complex globular structures with many cisternae or lobes. These structures are dotted with viral replication complexes and are often called "replication vesicles", "viroplasm" or "viral factories". Replication vesicle membranes are derived from

2538-533: Is one of many enzymes in the Calvin cycle . When Rubisco facilitates the attack of CO 2 at the C2 carbon of RuBP and subsequent bond cleavage between the C3 and C2 carbon, 2 molecules of glycerate-3-phosphate are formed. The conversion involves these steps: enolisation , carboxylation , hydration , C-C bond cleavage, and protonation . Substrates for RuBisCO are ribulose-1,5-bisphosphate and carbon dioxide (distinct from

2632-590: Is released into host cytoplasm. Potyviral RNA mimicks host mRNA : the 5' VPg protein shares functional similarities with the 5' cap and the 3' end is polyadenylated . VPg and its interactions with eIF4E and eIF4(iso)E allow the virus to utilise host cap-dependent translation machinery for its translation. Similarly to eukaryotic translation, the VPg-eIF4E interaction assembles the eIF4F complex around viral RNA. A number of weak internal ribosome entry sites (IRES) have been identified in many potyvirus species but it

2726-426: Is required because ribulose 1,5-bisphosphate (RuBP) binds more strongly to the active sites of RuBisCO when excess carbamate is present, preventing processes from moving forward. In the light, RuBisCO activase promotes the release of the inhibitory (or — in some views — storage) RuBP from the catalytic sites of RuBisCO. Activase is also required in some plants (e.g., tobacco and many beans) because, in darkness, RuBisCO

2820-466: Is required for cell-to-cell movement, the central hydrophobic transmembrane alpha helix , and the C terminal domain which is required for viral replication. P3N-PIPO (~25 kDa) is a dedicated movement protein which anchors the movement complex to the plasmodesma . It may also modulate the plasmodesmatal size exclusion limit by interacting with host proteins which sever plasmodesmatal actin filaments and reduce callose deposition. It interacts with both

2914-468: Is slow, fixing only 3-10 carbon dioxide molecules each second per molecule of enzyme. The reaction catalyzed by RuBisCO is, thus, the primary rate-limiting factor of the Calvin cycle during the day. Nevertheless, under most conditions, and when light is not otherwise limiting photosynthesis, the speed of RuBisCO responds positively to increasing carbon dioxide concentration. RuBisCO is usually only active during

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3008-476: Is the substrate, the products of the oxygenase reaction are phosphoglycolate and 3-phosphoglycerate. Phosphoglycolate is recycled through a sequence of reactions called photorespiration , which involves enzymes and cytochromes located in the mitochondria and peroxisomes (this is a case of metabolite repair ). In this process, two molecules of phosphoglycolate are converted to one molecule of carbon dioxide and one molecule of 3-phosphoglycerate, which can reenter

3102-664: The Retroviridae (e.g. HIV ), genome damage appears to be avoided during reverse transcription by strand switching, a form of recombination. Recombination occurs in the Coronaviridae (e.g. SARS ). Recombination in RNA viruses appears to be an adaptation for coping with genome damage. Recombination can also occur infrequently between +ssRNA viruses of the same species but of divergent lineages. The resulting recombinant viruses may sometimes cause an outbreak of infection in humans, as in

3196-480: The host cell's ribosomes . Positive-strand RNA viruses encode an RNA-dependent RNA polymerase (RdRp) which is used during replication of the genome to synthesize a negative-sense antigenome that is then used as a template to create a new positive-sense viral genome. Positive-strand RNA viruses are divided between the phyla Kitrinoviricota , Lenarviricota , and Pisuviricota (specifically classes Pisoniviricetes and Stelpavirictes ) all of which are in

3290-540: The 3-hydroxypropionate cycle , or the reverse Krebs cycle , these pathways are relatively small contributors to global carbon fixation compared to that catalyzed by RuBisCO. Phosphoenolpyruvate carboxylase , unlike RuBisCO, only temporarily fixes carbon. Reflecting its importance, RuBisCO is the most abundant protein in leaves , accounting for 50% of soluble leaf protein in C 3 plants (20–30% of total leaf nitrogen) and 30% of soluble leaf protein in C 4 plants (5–9% of total leaf nitrogen). Given its important role in

3384-544: The Calvin Cycle ) from taking place, since these reactions require CO 2 to pass by gas exchange through these openings. Evaporation through the upper side of a leaf is prevented by a layer of wax . Since RuBisCO is often rate-limiting for photosynthesis in plants, it may be possible to improve photosynthetic efficiency by modifying RuBisCO genes in plants to increase catalytic activity and/or decrease oxygenation rates. This could improve sequestration of CO 2 and be

3478-475: The Tobacco mosaic virus , potyviruses do not have a single movement protein but instead assemble a movement complex around the plasmodesma. This complex is primarily composed of three viral proteins: CI, CP, P3N-PIPO. Conical CI inclusions are anchored to plasmodesmata by P3N-PIPO during the early stages of potyvirus infection. This allows the inclusion to funnel either viral particles or viral RNA-CP complexes through

3572-421: The methionine salvage pathway . Later identifications found functionally divergent examples dispersed all over bacteria and archaea, as well as transitionary enzymes performing both RLP-type enolase and RuBisCO functions. It is now believed that the current RuBisCO evolved from a dimeric RLP ancestor, acquiring its carboxylase function first before further oligomerizing and then recruiting the small subunit to form

3666-424: The thylakoid membrane. The movement of protons into thylakoids is driven by light and is fundamental to ATP synthesis in chloroplasts (Further reading: Photosynthetic reaction centre ; Light-dependent reactions ) . To balance ion potential across the membrane, magnesium ions ( Mg ) move out of the thylakoids in response, increasing the concentration of magnesium in the stroma of the chloroplasts. RuBisCO has

3760-405: The "activating" carbon dioxide). RuBisCO also catalyses a reaction of ribulose-1,5-bisphosphate and molecular oxygen (O 2 ) instead of carbon dioxide (CO 2 ). Discriminating between the substrates CO 2 and O 2 is attributed to the differing interactions of the substrate's quadrupole moments and a high electrostatic field gradient . This gradient is established by the dimer form of

3854-424: The C 3 cycle was shown to be possible, and it was first achieved in 2019 through a synthetic biology approach. One avenue is to introduce RuBisCO variants with naturally high specificity values such as the ones from the red alga Galdieria partita into plants. This may improve the photosynthetic efficiency of crop plants, although possible negative impacts have yet to be studied. Advances in this area include

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3948-732: The C2-C3 bond to form one molecule of glycerate-3-phosphate and a negatively charged carboxylate. Stereo specific protonation of C2 of this carbanion results in another molecule of glycerate-3-phosphate. This step is thought to be facilitated by Lys175 or potentially the carbamylated Lys210. When carbon dioxide is the substrate, the product of the carboxylase reaction is an unstable six-carbon phosphorylated intermediate known as 3-keto-2-carboxyarabinitol-1,5-bisphosphate, which decays rapidly into two molecules of glycerate-3-phosphate . This product, also known as 3-phosphoglycerate, can be used to produce larger molecules such as glucose . When molecular oxygen

4042-503: The C3 carbon of RuBP to form a 2,3-enediolate. Carboxylation of the 2,3-enediolate results in the intermediate 3-keto-2-carboxyarabinitol-1,5-bisphosphate and Lys334 is positioned to facilitate the addition of the CO 2 substrate as it replaces the third Mg -coordinated water molecule and add directly to the enediol. No Michaelis complex is formed in this process. Hydration of this ketone results in an additional hydroxy group on C3, forming

4136-535: The Calvin cycle. Some of the phosphoglycolate entering this pathway can be retained by plants to produce other molecules such as glycine . At ambient levels of carbon dioxide and oxygen, the ratio of the reactions is about 4 to 1, which results in a net carbon dioxide fixation of only 3.5. Thus, the inability of the enzyme to prevent the reaction with oxygen greatly reduces the photosynthetic capacity of many plants. Some plants, many algae, and photosynthetic bacteria have overcome this limitation by devising means to increase

4230-559: The P3 protein, which is generated into a subset of transcripts by a +2 frameshift caused by a ribosome slippage mechanism at a conserved GA 6 repeat sequence. The resulting protein is called P3N-PIPO. A similar mechanism is thought to produce an alternative reading frame within the P1 cistron, named "pretty interesting sweet potato potyvirus ORF" (PISPO), in a number of sweet potato-infecting potyviruses including sweet potato feathery mottle virus . P1 (~33 kilo daltons (kDa) in molecular weight)

4324-491: The activation state of RuBisCO can be modulated in response to light intensity and, thus, the rate of formation of the ribulose 1,5-bisphosphate substrate. In cyanobacteria, inorganic phosphate (P i ) also participates in the co-ordinated regulation of photosynthesis: P i binds to the RuBisCO active site and to another site on the large chain where it can influence transitions between activated and less active conformations of

4418-456: The active site by the rotation of His335 to an alternate conformation. Mg is then coordinated by the His residues of the active site (His300, His302, His335), and is partially neutralized by the coordination of three water molecules and their conversion to OH. This coordination results in an unstable complex, but produces a favorable environment for the binding of Mg . Formation of the carbamate

4512-838: The aphid and typically only persist in the aphid for a few minutes. Certain potyviruses have been shown to alter the feeding patterns of their aphid vectors, which may manifest as longer periods of time spent on infected plants, reduced non-probing feeding time, and increased phloem sap ingestion. Seed and pollen transmission has been documented in certain potyvirus species, for instance in PVY and Turnip mosaic virus (TUMV) . Vegetative transmission by infected tubers or grafting material are of particular concern for certain agricultural crops, such as potato and fruit trees, respectively. Transmission can also occur by physical contact with infected plants or with contaminated tools, clothes, or even water. After entry, potyvirus particles are uncoated and genomic RNA

4606-478: The apparent descendants of leviviruses, which infect eukaryotes . The phylum is divided into four classes: Leviviricetes , which contains leviviruses and their relatives, Amabiliviricetes , which contains narnaviruses and their relatives, Howeltoviricetes , which contains mitoviruses and their relatives, and Miaviricetes , which contains botourmiaviruses and their relatives. Based on phylogenetic analysis of RdRp, all other RNA viruses are considered to comprise

4700-454: The binding sites. A total of eight large chains (= four dimers) and eight small chains assemble into a larger complex of about 540,000 Da. In some Pseudomonadota and dinoflagellates , enzymes consisting of only large subunits have been found. Magnesium ions ( Mg ) are needed for enzymatic activity. Correct positioning of Mg in the active site of the enzyme involves addition of an "activating" carbon dioxide molecule ( CO 2 ) to

4794-411: The biosphere, the genetic engineering of RuBisCO in crops is of continuing interest (see below ). In plants, algae , cyanobacteria , and phototrophic and chemoautotrophic Pseudomonadota (formerly proteobacteria), the enzyme usually consists of two types of protein subunit, called the large chain ( L , about 55,000 Da ) and the small chain ( S , about 13,000 Da). The large-chain gene ( rbcL )

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4888-739: The case of SARS and MERS. Positive-strand RNA viruses are common in plants. In tombusviruses and carmoviruses , RNA recombination occurs frequently during replication. The ability of the RNA-dependent RNA polymerase of these viruses to switch RNA templates suggests a copy choice model of RNA recombination that may be an adaptive mechanism for coping with damage in the viral genome. Other +ssRNA viruses of plants have also been reported to be capable of recombination, such as Brom mosaic bromovirus and Sindbis virus . Positive-strand RNA viruses are found in three phyla: Kitrinoviricota , Lenarviricota , and Pisuviricota , each of which are assigned to

4982-453: The cell's secretory pathway for viral replication. Numerous positive-strand RNA viruses can undergo genetic recombination when at least two viral genomes are present in the same host cell. The capability for recombination among +ssRNA virus pathogens of humans is common. RNA recombination appears to be a major driving force in determining genome architecture and the course of viral evolution among Picornaviridae (e.g. poliovirus). In

5076-595: The cleavage sites of the polyprotein. The only exceptions are the self-cleavages of P1 and HC-Pro. The high degree of cleavage sequence specificity and conservation has made NIa-Pro (often that of Tobacco etch virus ) a valuable tool in biotechnology, especially in applications which require removing affinity tags from recombinant proteins after affinity purification . NIa-Pro has also shown to exhibit sequence-independent DNase activity and to interfere with host DNA methylation suggesting that NIa and/or NIa-Pro are altering in host gene expression . Potyviral NIa-Pro shares

5170-687: The concentration of carbon dioxide around the enzyme, including C 4 carbon fixation , crassulacean acid metabolism , and the use of pyrenoid . Rubisco side activities can lead to useless or inhibitory by-products. Important inhibitory by-products include xylulose 1,5-bisphosphate and glycero-2,3-pentodiulose 1,5-bisphosphate , both caused by "misfires" halfway in the enolisation-carboxylation reaction. In higher plants, this process causes RuBisCO self-inhibition, which can be triggered by saturating CO 2 and RuBP concentrations and solved by Rubisco activase (see below). Some enzymes can carry out thousands of chemical reactions each second. However, RuBisCO

5264-420: The consumption of ATP . This reaction is inhibited by the presence of ADP , and, thus, activase activity depends on the ratio of these compounds in the chloroplast stroma. Furthermore, in most plants, the sensitivity of activase to the ratio of ATP/ADP is modified by the stromal reduction/oxidation ( redox ) state through another small regulatory protein, thioredoxin . In this manner, the activity of activase and

5358-519: The coronaviruses and rhinoviruses that cause the common cold . Positive-strand RNA virus genomes usually contain relatively few genes, usually between three and ten, including an RNA-dependent RNA polymerase. Coronaviruses have the largest known RNA genomes, between 27 and 32 kilobases in length, and likely possess replication proofreading mechanisms in the form of an exoribonuclease within nonstructural protein nsp14. Positive-strand RNA viruses have genetic material that can function both as

5452-428: The day, as ribulose 1,5-bisphosphate is not regenerated in the dark. This is due to the regulation of several other enzymes in the Calvin cycle. In addition, the activity of RuBisCO is coordinated with that of the other enzymes of the Calvin cycle in several other ways: Upon illumination of the chloroplasts, the pH of the stroma rises from 7.0 to 8.0 because of the proton (hydrogen ion, H ) gradient created across

5546-557: The detection of various proteinaceous inclusions in infected plant cells. These may appear as crystals in either the cytoplasm or in the nucleus, as amorphous X-bodies, membranous bodies, viroplasms or pinwheels. The inclusions may or may not (depending on the species) contain virions. These inclusions can be seen by light microscopy in leaf strips of infected plant tissue stained with Orange-Green (protein stain) but not Azure A (nucleic acid stain). Modern detection methods rely primarily on reverse transcription PCR . Potyvirus contains

5640-505: The enzyme must be closed off, allowing the active site to be isolated from the solvent and lowering the dielectric constant . This isolation has a significant entropic cost, and results in the poor turnover rate. Carbamylation of the ε-amino group of Lys210 is stabilized by coordination with the Mg . This reaction involves binding of the carboxylate termini of Asp203 and Glu204 to the Mg ion. The substrate RuBP binds Mg displacing two of

5734-444: The enzyme. In this way, activation of bacterial RuBisCO might be particularly sensitive to P i levels, which might cause it to act in a similar way to how RuBisCO activase functions in higher plants. Since carbon dioxide and oxygen compete at the active site of RuBisCO, carbon fixation by RuBisCO can be enhanced by increasing the carbon dioxide level in the compartment containing RuBisCO ( chloroplast stroma ). Several times during

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5828-497: The evolution of plants, mechanisms have evolved for increasing the level of carbon dioxide in the stroma (see C 4 carbon fixation ). The use of oxygen as a substrate appears to be a puzzling process, since it seems to throw away captured energy. However, it may be a mechanism for preventing carbohydrate overload during periods of high light flux. This weakness in the enzyme is the cause of photorespiration , such that healthy leaves in bright light may have zero net carbon fixation when

5922-446: The familiar modern enzyme. The small subunit probably first evolved in anaerobic and thermophilic organisms, where it enabled RuBisCO to catalyze its reaction at higher temperatures. In addition to its effect on stabilizing catalysis, it enabled the evolution of higher specificities for CO 2 over O 2 by modulating the effect that substitutions within RuBisCO have on enzymatic function. Substitutions that do not have an effect without

6016-550: The following species: A further four viruses were previously classified as species in this genus but were abolished due to lack of genetic sequence information: Potyviruses were further divided into the PVY, SCMV, BYMV, BCMV species groups in 1992. Gibbs and Ohshima 2010 produced a more extensive molecular phylogeny with the same four, but also several new groups: the BtMV, ChVMV, DaMV, OYDV, PRSV, TEV, and TuMV. Contains 16 species including

6110-465: The interactions between replication vesicles and movement complex proteins which may allow replication vesicles to be recruited to the movement complex for efficient intercellular movement. P3 also interacts with large subunit of the ribulose-1,5-bisphosphate carboxylase/oxygenase . CI (~71 kDa) is an RNA helicase with ATPase activity. Its most unusual property is its ability to form large and highly symmetrical conical and cylindrical inclusions with

6204-590: The kingdom Orthornavirae and realm Riboviria . They are monophyletic and descended from a common RNA virus ancestor. In the Baltimore classification system, +ssRNA viruses belong to Group IV. Positive-sense RNA viruses include pathogens such as the Hepatitis C virus , West Nile virus , dengue virus , and the MERS , SARS , and SARS-CoV-2 coronaviruses , as well as less clinically serious pathogens such as

6298-423: The kingdom Orthornavirae in the realm Riboviria . In the Baltimore classification system, which groups viruses together based on their manner of mRNA synthesis, +ssRNA viruses are group IV. The first +ssRNA phylum is Kitrinoviricota . The phylum contains what have been referred to as the " alphavirus supergroup" and " flavivirus supergroup" along with various other short-genome viruses. Four classes in

6392-468: The large and small subunits of the ribulose-1,5-bisphosphate carboxylase/oxygenase. CP (~30 - 35 kDa) is the capsid protein. It has two terminal domains which are disordered and exposed at the surface of the virion. The central core domain contains an RNA-binding pocket which binds to viral RNA. The structure of the capsid protein is highly conserved in potyviruses, though there is a relatively high degree of sequence variability. In addition to encapsidating

6486-407: The large subunit of RuBisCO has been widely used as an appropriate locus for analysis of phylogenetics in plant taxonomy . Non-carbon-fixing proteins similar to RuBisCO, termed RuBisCO-like proteins (RLPs), are also found in the wild in organisms as common as Bacillus subtilis . This bacterium has a rbcL-like protein with a 2,3-diketo-5-methylthiopentyl-1-phosphate enolase function, part of

6580-462: The level of RNA interference suppression during infection. HC-Pro is also involved in aphid transmission. Though the exact mechanism is unknown, HC-Pro has been proposed to attach to host aphid mouth parts through its N-terminal zinc finger -like domain and anchor virions through its interactions with the capsid protein. P3 (~41 kDa) is a membrane protein which is required for viral replication and accumulates in viral replication vesicles. It mediates

6674-451: The membranes of a variety of organelles —often the rough endoplasmic reticulum , but also including membranes derived from mitochondria , vacuoles , the Golgi apparatus , chloroplasts , peroxisomes , plasma membranes , autophagosomal membranes , and novel cytoplasmic compartments. The replication of the positive-sense RNA genome proceeds through double-stranded RNA intermediates, and

6768-516: The minimally active RuBisCO, which with its two components provides a combination of oppositely charged domains required for the enzyme's interaction with O 2 and CO 2 . These conditions help explain the low turnover rate found in RuBisCO: In order to increase the strength of the electric field necessary for sufficient interaction with the substrates’ quadrupole moments , the C- and N- terminal segments of

6862-500: The mutant plants grew more slowly than wild-type. A recent theory explores the trade-off between the relative specificity (i.e., ability to favour CO 2 fixation over O 2 incorporation, which leads to the energy-wasteful process of photorespiration ) and the rate at which product is formed. The authors conclude that RuBisCO may actually have evolved to reach a point of 'near-perfection' in many plants (with widely varying substrate availabilities and environmental conditions), reaching

6956-464: The newly-evolved enzyme was found to have further developed a series of stabilizing mutations. While RuBisCO has always been accumulating new mutations, most of these mutations that have survived have not had significant effects on protein stability. The destabilizing C 4 mutations on RuBisCO has been sustained by environmental pressures such as low CO 2 concentrations, requiring a sacrifice of stability for new adaptive functions. The term "RuBisCO"

7050-464: The nuclear-encoded RbcS subunits, which are typically imported into chloroplasts as unfolded proteins. Furthermore, sufficient expression and interaction with Rubisco activase are major challenges as well. One successful method for expression of Rubisco in E. coli involves the co-expression of multiple chloroplast chaperones, though this has only been shown for Arabidopsis thaliana Rubisco. Due to its high abundance in plants (generally 40% of

7144-483: The nucleocapsid is helical with a pitch of 3.4-3.5 nm. The genome is a linear, positive-sense , single-stranded RNA ranging in size from 9,000 to 12,000 nucleotide bases . Most potyviruses have non-segmented genomes, though a number of species are bipartite. The typical base compositions of some of the most common, non-recombinant strains of the type species, PVY, range between ~23.4-23.8 % G; ~31-31.6 % A; ~18.2-18.8 % C; and ~26.5-26.8 % U. In

7238-451: The phylum are recognized: Alsuviricetes , the alphavirus supergroup, which contains a large number of plant viruses and arthropod viruses; Flasuviricetes , which contains flaviviruses, Magsaviricetes , which contains nodaviruses and sinhaliviruses ; and Tolucaviricetes , which primarily contains plant viruses. Lenarviricota is the second +ssRNA phylum. It contains the class Leviviricetes , which infect prokaryotes , and

7332-497: The plasmodesma. Replication vesicles are also recruited to the movement complex, suggesting that replication and movement are coupled. Replication vesicles are recruited by P3N-PIPO, which interacts with both CI and P3 through the shared P3N-domain. P3's interaction with 6K2 allows replication vesicles to be tethered to the movement complex. Potyviruses evolved between 6,600 and 7,250 years ago. They appear to have evolved in southwest Eurasia or north Africa . The estimated mutation rate

7426-528: The purpose of replication in these membranous invaginations may be the avoidance of cellular response to the presence of dsRNA. In many cases subgenomic RNAs are also created during replication. After infection, the entirety of the host cell's translation machinery may be diverted to the production of viral proteins as a result of the very high affinity for ribosomes by the viral genome's internal ribosome entry site (IRES) elements; in some viruses, such as poliovirus and rhinoviruses , normal protein synthesis

7520-407: The rate of carbon fixation. In general, site-directed mutagenesis of RuBisCO has been mostly unsuccessful, though mutated forms of the protein have been achieved in tobacco plants with subunit C 4 species, and a RuBisCO with more C 4 -like kinetic characteristics have been attained in rice via nuclear transformation. Robust and reliable engineering for yield of RuBisCO and other enzymes in

7614-448: The ratio of O 2 to CO 2 available to RuBisCO shifts too far towards oxygen. This phenomenon is primarily temperature-dependent: high temperatures can decrease the concentration of CO 2 dissolved in the moisture of leaf tissues. This phenomenon is also related to water stress : since plant leaves are evaporatively cooled, limited water causes high leaf temperatures. C 4 plants use the enzyme PEP carboxylase initially, which has

7708-482: The release of CA1P from the catalytic sites. After the CA1P is released from RuBisCO, it is rapidly converted to a non-inhibitory form by a light-activated CA1P-phosphatase . Even without these strong inhibitors, once every several hundred reactions, the normal reactions with carbon dioxide or oxygen are not completed; other inhibitory substrate analogs are still formed in the active site. Once again, RuBisCO activase can promote

7802-400: The release of these analogs from the catalytic sites and maintain the enzyme in a catalytically active form. However, at high temperatures, RuBisCO activase aggregates and can no longer activate RuBisCO. This contributes to the decreased carboxylating capacity observed during heat stress. The removal of the inhibitory RuBP, CA1P, and the other inhibitory substrate analogs by activase requires

7896-628: The replacement of the tobacco enzyme with that of the purple photosynthetic bacterium Rhodospirillum rubrum . In 2014, two transplastomic tobacco lines with functional RuBisCO from the cyanobacterium Synechococcus elongatus PCC7942 (Se7942) were created by replacing the RuBisCO with the large and small subunit genes of the Se7942 enzyme, in combination with either the corresponding Se7942 assembly chaperone, RbcX, or an internal carboxysomal protein, CcmM35. Both mutants had increased CO 2 fixation rates when measured as carbon molecules per RuBisCO. However,

7990-399: The small subunit suddenly become beneficial when it is bound. Furthermore, the small subunit enabled the accumulation of substitutions that are only tolerated in its presence. Accumulation of such substitutions leads to a strict dependence on the small subunit, which is observed in extant Rubiscos that bind a small subunit. With the mass convergent evolution of the C 4 -fixation pathway in

8084-726: The species with a monopartite genome, a genome-linked VPg protein is covalently bound to the 5' end and the 3' end is polyadenylated. The genome encodes a single open reading frame (ORF) expressed as a 350 kDa polyprotein precursor. This polyprotein is processed into ten smaller proteins: protein 1 protease (P1-Pro), helper component protease (HC-Pro), protein 3 (P3), cylindrical inclusion (CI), viral protein genome-linked (Vpg), nuclear inclusion A (NIa), nuclear inclusion B (NIb), capsid protein (CP) and two small putative proteins known as 6K1 and 6K2. The P3 cistron also contains an overlapping reading frame called "Pretty interesting Potyviridae ORF " (PIPO). PIPO codes for an alternative C-terminus to

8178-419: The subfamily Aphidinae (genera Macrosiphum and Myzus ). The genus contains 190 species and potyviruses account for about thirty percent of all currently known plant viruses . The virion is non-enveloped with a flexuous and filamentous nucleocapsid , 680 to 900 nanometers (nm) long and is 11–20 nm in diameter. The nucleocapsid contains around 2000 copies of the capsid protein. The symmetry of

8272-461: The three aquo ligands. Enolisation of RuBP is the conversion of the keto tautomer of RuBP to an enediol(ate). Enolisation is initiated by deprotonation at C3. The enzyme base in this step has been debated, but the steric constraints observed in crystal structures have made Lys210 the most likely candidate. Specifically, the carbamate oxygen on Lys210 that is not coordinated with the Mg ion deprotonates

8366-468: The total protein content), RuBisCO often impedes analysis of important signaling proteins such as transcription factors , kinases , and regulatory proteins found in lower abundance (10-100 molecules per cell) within plants. For example, using mass spectrometry on plant protein mixtures would result in multiple intense RuBisCO subunit peaks that interfere and hide those of other proteins. Recently, one efficient method for precipitating out RuBisCO involves

8460-486: The type species of the genus (potato virus Y). The primary hosts are: Nine Solanaceae , three Amaranthus , three Asteraceae , one Lilium , and one Amaryllis . Positive-strand RNA virus Positive-strand RNA viruses ( +ssRNA viruses ) are a group of related viruses that have positive-sense , single-stranded genomes made of ribonucleic acid . The positive-sense genome can act as messenger RNA (mRNA) and can be directly translated into viral proteins by

8554-459: The usage of protamine sulfate solution. Other existing methods for depleting RuBisCO and studying lower abundance proteins include fractionation techniques with calcium and phytate, gel electrophoresis with polyethylene glycol, affinity chromatography , and aggregation using DTT , though these methods are more time-consuming and less efficient when compared to protamine sulfate precipitation. The chloroplast gene rbcL , which codes for

8648-519: The virion, CP core domain is required for intercellular movement and contributes to seed transmission. Certain atypical potyviruses code for additional proteins or protein domains, such as P1-PISPO, Alkylation B (AlkB) , and inosine triphosphate pyrophosphatase (known as ITPase or HAM1). Such anomalies are often situated in the hypervariable P1-HC-Pro region. Most potyviruses are transmitted by aphids as they probe plant tissues with their stylet during feeding. They do not circulate or multiply within

8742-400: Was coined humorously in 1979, by David Eisenberg at a seminar honouring the retirement of the early, prominent RuBisCO researcher, Sam Wildman , and also alluded to the snack food trade name " Nabisco " in reference to Wildman's attempts to create an edible protein supplement from tobacco leaves. The capitalization of the name has been long debated. It can be capitalized for each letter of

8836-440: Was constrained by the trade-off between stability and activity brought about by the series of necessary mutations for C 4 RuBisCO. Moreover, in order to sustain the destabilizing mutations, the evolution to C 4 RuBisCO was preceded by a period in which mutations granted the enzyme increased stability, establishing a buffer to sustain and maintain the mutations required for C 4 RuBisCO. To assist with this buffering process,

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