key: cord-0777307-bppr91ve authors: Hassan, Sk. Sarif; Choudhury, Pabitra Pal; Uversky, Vladimir N.; Dayhoff, Guy W.; Aljabali, Alaa A. A.; Uhal, Bruce D.; Lundstrom, Kenneth; Rezaei, Nima; Seyran, Murat; Pizzol, Damiano; Adadi, Parise; Lal, Amos; Soares, Antonio; Abd El-Aziz, Tarek Mohamed; Kandimalla, Ramesh; Tambuwala, Murtaza; Azad, Gajendra Kumar; Sherchan, Samendra P.; Baetas-da-Cruz, Wagner; Takayama, Kazuo; Serrano-Aroca, Ángel; Chauhan, Gaurav; Palu, Giorgio; Brufsky, Adam M. title: Variability of Accessory Proteins Rules the SARS-CoV-2 Pathogenicity date: 2020-11-08 journal: bioRxiv DOI: 10.1101/2020.11.06.372227 sha: 36d3dabfddb423c0622732862b8000fed2df840f doc_id: 777307 cord_uid: bppr91ve The coronavirus disease 2019 (COVID-19) is caused by the Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) which is pandemic with an estimated fatality rate less than 1% is ongoing. SARS-CoV-2 accessory proteins ORF3a, ORF6, ORF7a, ORF7b, ORF8, and ORF10 with putative functions to manipulate host immune mechanisms such as interferons, immune signaling receptor NLRP3 (NOD-, LRR-, and pyrin domain-containing 3) inflammasome, inflammatory cytokines such as interleukin 1β (IL-1β) are critical in COVID-19 pathology. Outspread variations of each of the six accessory proteins of all complete proteomes (available as of October 26, 2020, in the National Center for Biotechnology Information depository) of SARS-CoV-2, were observed across six continents. Across all continents, the decreasing order of percentage of unique variations in the accessory proteins was found to be ORF3a>ORF8>ORF7a>ORF6>ORF10>ORF7b. The highest and lowest unique variations of ORF3a were observed in South America and Oceania, respectively. This finding suggests that the wide variations of accessory proteins seem to govern the pathogenicity of SARS-CoV-2, and consequently, certain propositions and recommendations can be made in the public interest. SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus-2), the causative agent for The coronavirus disease 2019 , the pandemic is ongoing with the estimated fatality rate less than 1% [1] . However, the World Health Organi- had 8 open reading frame (ORF) 3a, 3b, 6, 7a, 7b, 8a, 8b, and 9b which were suggested to have more intrinsic and secondary 10 other than having primary roles in the cell cycle and cellular entry [4, 5] . For instance, the ORFs are transcripted through the second phase of replication by (+) subgenomic messenger RNAs that were transcripted by the viral replication transcription complex negative-sense viral RNA coded in the initial stages of SARS-CoV-2 infection [6] . Thus, due to their intrinsic nature accessory proteins are not positive-selection sites such as extrinsic and primary functional Spike protein receptor-binding domain or protease cleavage sites [7] . Since, clinical SARS-CoV-2 isolates had a high-frequency non-synonymous mutation, 15 D614G, in their S protein, which increased host cell entry via ACE2 and Transmembrane Protease Serine 2 (TMPRSS2) [8] . Therefore, due to the intrinsic nature and secondary order in the viral transcription, we can expect less selective pressure and mutations on accessory proteins to reach high-frequency in the population is less expected. Thus, despite the 19 to 89 years of estimated genomic divergence between SARS-CoV-2 and Ratg13, the sequence identity on accessory proteins ORF3, ORF6, ORF7a, ORF7b, ORF8, ORF10 had 98.5, 100, 97.5, 97.6, 95, and 100%, respectively are very high or identical which 20 is indicating somehow the direct ancestor of SARS-CoV-2 had been exposed to almost no selective pressure to manipulate its intermediate host immunity for many years until the primary Human infection in Wuhan ( Fig.1 to Fig.6) [2]. SARS-CoV-2 and SARS-CoV accessory proteins have differences such as putative protein ORF10 not present in SARS-CoV and the ORF3b and ORF9b is not present in SARS-CoV-2 [9, 10] . Very little is known about the functions of accessory proteins of SARS-CoV-2. The known essential features of the six accessory proteins are summarized below. 25 ORF3a Protein: The ORF3a is the 275 amino acids long largest accessory protein among the accessory proteins coded by the SARS-CoV-2, has 72.4% sequence identity with SARS-CoV ORF3a protein and has 98.5% sequence identity with BatCoV Ratg13 ORF3a protein [11, 12] (Fig.1 ). ORF3a is involved in virulence, infectivity, ion channel activity, morphogenesis, and virus release [13] . In SARS-CoV ORF3a was a multifunctional protein, co-localized with its protein binding regions with E, M, and S proteins in viral 30 assembly formed homo-tetrameric complex as potassium-ion channel on host membrane [5] . In SARS-CoV-2, the ion-channel proteins (viroporins) function of ORF3a in addition to other proteins such as protein E, and ORF8a is critical in CoVs tissue inflammation [6] . Viroporins mediated lysosomal disruption and ion-redistribution activates innate immune signaling receptor NLRP3 (NOD-, LRR-, and pyrin domain-containing 3) inflammasome that leads to the expression of inflammatory cytokines such as interleukin 1β (IL-1β), IL-6, and tumor necrosis factor (TNF), causing tissue inflammation during respiratory illness 35 [6] . From another pathway, ORF3a with its protein binding domains interacts with TNF receptor-associated factor (TRAF3) protein, which leads ASC ubiquitination, and caspase 1 activation, and IL-1β maturation [14] . Additionally, ORF3a and ORF7a in combination with E, S, Nsp1 protein, and MAPK pathway proteins (MAPK8, MAPK14, and MAP3K7) trigger proinflammatory cytokine signaling transcription factors such as STAT1, STAT2, IRF9, and NFKB1 [6] . Another SARS-CoV-2 ORF3a protein interacts with heme oxygenase-1 (HMOX1) that has a role in heme catabolism and the anti-inflammatory 40 system [6] . SARS-CoV-2 either triggers viral dissemination or suppresses continued viral replication of the apoptosis or programmed cell death [6] . In SARS-CoV ORF3a E and M protein, ion channel activity interferes with apoptotic pathways [11] . ORF6 Protein: SARS-CoV-2 ORF6 is a 61 amino acid long membrane-associated interferon (IFN) antagonist protein suppresses the expression of co-transfected expression constructs and its subcellular localization to vesicular structures that 45 has 68.9% sequence identity, with SARS-CoV ORF6 protein and has 100% sequence identity, with BatCoV Ratg13 ORF6 protein [5] (Fig.2) . ORF6 interacts with the karyopherin import complex that limits the movement of transcription factors STAT1 which down-regulates the IFN pathway [5] . In SARS-CoV ORF6 and ORF3a, in association with other proteins such as M, Nsp1 with Nsp3 inhibit IRF3 signalling and repress interferon expression and stimulate the degradation of IFNAR1 and STAT1 50 [6] . ORF6 interacts with the nsp8 protein coded by SARS-CoV-2 and it can increase infection during early infection at a low multiplicity with increase in RNA polymerase activity [15] . It is reported that ORF6 and ORF8 can inhibit the type-I interferon signaling pathway [15] . ORF6 protein with lysosomal targeting motif (YSEL) and diacidic motif (DDEE) induces intracellular membrane rearrangements resulting in a vesicular population and endosomal internalization of viral protein into the infected cells, increasing replication [16] . ORF7a 121 aa coding type I transmembrane protein interacts with SARS-CoV-2 structural proteins M, E, and S, which are essential for viral assembly, and hence ORF7a is involved in the viral replication cycle and virion-associated ORF7a protein may function during early infection that has 85.2% Sequence identity with SARS-CoV ORF7a protein and has 97.5% sequence identity, with BatCoV Ratg13 ORF7a protein [5] (Fig.3 ). 60 Figure 3 : ClustalW alignment of SARS-CoV-2 (NCBI GenBank ID BCA87366.1) and Ratg13 (NCBI GenBank ID MN996532.2, translated 5'3' frame 2) ORF7a proteins shows 97.5% sequence identity, despite up to 89 years of genetic diversion. ORF7a interacts with SARS-CoV-2 structural proteins: membrane (M), envelope (E), and spike (S), which are essential for viral assembly, and hence ORF7a is involved in the viral replication cycle and virion-associated ORF7a protein may function during early infection [17, 18] . ORF7a leads the activation of pro-inflammatory cytokines and chemokines, such as IL-8 and RANTES [5] . SARS-CoV ORF7a in combination with E protein activate apoptosis by suppressing anti-apoptotic protein [6] . ORF7b is a 43 aa coding protein found in association with intracellular virus particles and also in purified virions inside the Golgi compartment that has an 85.4% sequence identity with SARS-CoV ORF7b protein and has 97.6% sequence identity, with BatCoV Ratg13 ORF7a protein [5] (Fig.4) . ORF7b is found in association with intracellular virus particles and also in purified virions. Till date, there is very little experimental evidence to support a role for ORF7a or ORF7b in the replication of SARS-CoV-2 [19] . ORF8 of SARS-CoV-2 interacts with major histocompatibility complex (MHC) class-I molecules and down-regulates their 75 surface expression significantly on various cell types [21] . It has been reported earlier that inhibition of ORF8 function could be a strategy to improve the special immune surveillance and accelerate the eradication of SARS-CoV-2 in vivo [22] . The accessory protein 38 aa coding protein ORF10 has been reported to be unique for SARS-CoV-2 containing eleven cytotoxic T lymphocyte (CTL) epitopes of nine amino acids in length each, across various human leukocyte antigen (HLA) subtypes [23, 24] . ORF10 negatively affects the antiviral protein degradation process through its interaction 80 with the Cul2 ubiquitin ligase complex [6] . SARS-CoV does not have ORF10 protein but SARS-COV-2 ORF10 and Ratg13 ORF10 has 97.3% sequence identity [25] (Fig.6 ). The objectives of the present study were to depict the unique variability of all accessory proteins and their possible contributions to virus pathogenicity. Sequences for all the accessory proteins ORF3a, ORF6, ORF7a, ORF7b, ORF8, and ORF10 were downloaded (on October, 20, 2020) from the complete SARS-CoV-2 proteomes on the National Center for Biotechnology Information (NCBI) database (http://www.ncbi.nlm.nih.gov/) ( Table 1) . Note that all partial accessory proteins and sequences with ambiguous amino acids were excluded from the present study. Furthermore, the unique accessory protein sequences were extracted for each continent. The unique protein accessions were 90 renamed for each accessory protein as S1, S2, . . . etc., as shown in the Supplementary Tables (7-13). There were 510, 72, 158, 37, 190, and 44 unique accessory proteins ORF3a, ORF6, ORF7a, ORF7b, ORF8, and ORF10, respectively, available. For each continent, ranges and names of sequences are presented in Table 2 . Africa S1 to S7 S1 to S3 S1 to S6 S1 to S2 S1 to S5 S1 Asia S8 to S85 S4 to S13 S7 to S25 S3 to S9 S6 to S31 S2 to S8 Europe S86 to S115 S14 to S19 S26 S10 to S11 S32 to S41 S9 to S12 North America S116 to S442 S20 to S58 S27 to S126 S12 to S30 S42 to S165 S13 to S36 Oceania S443 to S495 S59 to S69 S127 to S153 S31 to S36 S166 to S186 S37 to S42 South America S496 to S510 S70 to S72 S154 to S158 S37 S187 to S190 S43 to S44 Per-residue disorder distribution within the amino acid sequences of SARS-CoV-2 accessory proteins ORF3a, ORF6, ORF7a, ORF7b, ORF8 and ORF10 and their natural variants was evaluated by PONDR® VSL2, which is one of the more accurate standalone disorder predictors [26, 27, 28, 29] . The per-residue disorder predisposition scores are on a scale from 0 to 1, where values of 0 indicate fully ordered residues, and values of 1 indicate fully disordered residues. Values above the threshold of 0.5 are considered disordered residues, whereas residues with disorder scores between 0.25 and 0.5 are considered 100 highly flexible, and residues with disorder scores between 0.1 and 0.25 are taken as moderately flexible. For every continent, the total number of accessory proteins and the total number of unique sequences with respective percentages are presented in Fig.7 . In summary for all six continents, the total number of unique accessory proteins ORF3a, ORF6, ORF7a, ORF7b, ORF8, and ORF10 sequences are 419, 55, 122, 26, 147, and 32, respectively (Supplementary Fig.11 ). The percentage of each accessory protein across the six continents are presented as bar diagrams in Fig.8 . In addition, the percentage of unique accessory proteins among all unique sequences obtained across the six continents are represented as bar diagrams in Fig.9 . Following continent-wise, lists of identical sequences for each accessory protein were presented (Fig.10 ). The following observations were made for each accessory protein based on Fig.10 : ORF3a: Note that, the mutations described below were determined based on the Wuhan ORF3a sequence (YP 009724391). There were only two ORF3a sequences (marked in red font), S2 (with reference to Africa, QOI60359) and S5 (with 135 reference to Africa, QOI60335) which were present on all six continents. Note that the S2 (Africa-ORF3a) was identical with ORF3a (YP 009724391) from Wuhan, China. The other sequence S5 is different from ORF3a (YP 009724391) by one missense mutation Q57H, which was a strain determining mutation [30] . It is found that the ORF3a sequence S54 (Asia: QKK14624) possesses the single T175I mutation and is present on all continents except in Africa. The ORF3a sequences S62 (Asia: QMJ01306) and S63 (Asia: QJQ04482) possessed a single mutation each G251V and G196V, respectively with respect 140 to Wuhan ORF3a (YP 009724391). These two sequences were present in Asia, Europe, North America, Oceania, and South America. The ORF3a sequence S4 (Africa: QLQ87565) has the single S171L mutation found on four continents excluding were 57 unique ORF3a variants detected only on two continents as listed in Table 3 : ORF7b: Here all mutations are accounted based on the Wuhan ORF7b sequence (YP 009725318). The sequence S2 (ORF7b, Africa) (identical to Wuhan ORF7b (YP 009725318)) was found on all the six continents. It is found that only the C41F mutation was present in S8 (ORF7b, Asia) which appeared in Asia, North America, and Oceania. The sequence S1 (ORF7b, Africa) had the single mutation S5L and it was present in Africa and Asia. Sequence S5 (ORF7b, Asia) had the mutation S31L and this sequences was found on the two continents, Asia and North America 175 only. L32F occurred in the sequence S10 (ORF7b, Europe) which was present on the continents Europe and North America. Furthermore, ORF7b Sequence S13 had the mutation L4F and this sequence was found on North America and Oceania. ORF8: Mutations described below are determined with reference to the Wuhan ORF7b sequence (YP 009724396). It is observed that, the Wuhan ORF8 YP 009724396 sequence was found on every continent. Also, there was another sequence which is also present in every continent, having the single mutation L84S. The V62L, a single mutation was observed 180 in the sequence S2 (ORF8, Africa) which was found on all continents except South America, whereas ORF8 sequence S38 (Europe) possessed the single mutation A65S and the sequence was found in North America, Oceania, and South America. Further, V62L and L84S two mutations were observed in S12 (ORF8, Asia) and S12 appeared in Asia, North America, and Oceania. Sequence S15 (ORF8, Asia) got a mutation S67F and it was found in Asia, North America, and Oceania. ORF8 sequence S24 (Asia) possessed a single mutation A65V and the sequence was found in Asia, North America, and Oceania. The Wuhan ORF10 (YP 009725255) became identical with S1 (ORF10, Africa) and it was found on every continent. ORF10 sequence S6 (ORF10, Asia) had the mutation L37F and the sequence was present on North America, and Oceania only. The only mutation V30L was found in ORF10 sequence S10 (Europe) which appeared in Europe, North America, and Oceania. The sequence S9 (ORF10, Europe) had the mutation S23F and it was found in Europe and North America. Also, 190 the mutation D31Y appeared in S12 (ORF10, Europe) which was found in Europe and North America only. Here certain basic descriptive statistics (mean, variance, lower bound, upper bound, and range) were employed to describe the variability of the percentage of intrinsic protein disordered residues (IPD), molecular weight (MW), and isoelectric point (IP) of all the unique variants of all accessory proteins ( Table 6 ). The zigzag behavior of the plots of IPD, MW, and IP 195 depicts the wide variability of each accessory protein-variant ( Supplementary Fig.12-Fig.15 ). The following observations were made based on Table 6 . The amount total dispersions (based on range) of the percentage of IPD and MW of ORF6 variants turned out to be highest whereas the highest amount of total dispersion of IP was observed for ORF10. The smallest amount of total dispersions of the percentage of IPD, MW, and IP were found for ORF3a, ORF10, and ORF7b, respectively. The large value of range 200 and variance of the MW of the unique ORF3a, ORF7a, ORF8, and ORF10 variants imply the wide variability of each set of ORF3a, ORF7a, ORF8, and ORF10 though range and variance of IPD and IP were not much widely spread. In case of unique variance of ORF6, the range and variance of MW and percentage of IPD were found to be large which implied the wide quantitative differences among the unique ORF6 variants. Furthermore, moderately high range and variance associated with the percentage of IPD and MW of ORF7a variants imply its moderate variability. In line with the previously reported data, Fig.14 and Table 6 show that all SARS-CoV-2 accessory proteins contain different levels of intrinsic disorder. Furthermore, this analysis revealed that intrinsic disorder predispositions can vary significantly between the natural variants of each individual accessory protein. Importantly, the largest mutation-induced variability is observed within the disordered or flexible regions of these proteins (i.e., regions characterized by the predicted disorder scores exceeding the 0.5 threshold and regions with disorder scores between 0.25 and 0.5). This is an important observation 210 suggesting that natural variability of SARS-CoV-2 accessory proteins is shaping their structural flexibility. SARS-CoV-2 is the first HuCoVs with pandemic capacity due to its highly contagious nature deriving from the structural differences in its spike protein such as flat sialic acid binding domain, tight binding to its entry ACE2 receptor and capacity to be cleaved by furin protease [31] . However, based on the estimated infection number close to one billion by WHO, SARS-215 CoV-2 highly contagious but relatively a weak viral pathogen considering the overall of infection number has severe infections associated with the multiple organ dysfunctions [6] . This relatively weak pathological feature of SARS-CoV-2 could be related to the accessory proteins modulating host immunity as described above. Based on the dynamic and various mutations on accessory protein variants, SARS-CoV-2 after diverging with BatCoV cofactor proteins nsp10, nsp13, and nsp16 [32] . Muller's ratchet or also called as rachet effect explains the extinctive effect of high mutation rates of asexual organisms such as viruses [33] . Therefore, SARS-CoV-2 is repairing its mutations to preserve its genomic stability since a mutation can lead to pathological fitness losses or viral extinction [33] . However, there is a 225 balance governed by genomic repair mechanisms such as nsp14 and viruses that require a certain degree of mutations to gain novel traits such as emergence transmission in zoonotic hosts [33] . For instance, SARS-CoV mutations, a 29-nucleotide deletion of ORF8, was associated with the less pathogenic strain [33] . Similarly, SARS-CoV-2 variants with a 382-nucleotide deletion, ORF8 had mild symptoms and did not require supplemental oxygen [33] . Furthermore, only one variant (identical to the Wuhan sequence (NC 045512) of each of the accessory proteins of ORF6, ORF7a, ORF7b, and ORF10 were present on all continents. Furthermore, it was observed that only two variants of ORF3a differed by a single mutation (Q57H, a clade/strain determining [30] ) were found on all six continents. Also, in ORF8, only two unique variants (differed by a strain determining single mutation L84S) appeared on all continents. So, the maximally intersecting family of variations across all accessory proteins has turned out to be the smallest. These findings confirmed that the other variants of all the accessory proteins were due to demographic, environmental constraints. It was found that most of the unique variants of accessory proteins differed from the respective Wuhan accessory proteins by a single mutation, although basic descriptive statistics as found in section 3.1, unfolded their respective wide variability. Note that new variants of each accessory protein have been found in recent days and continue to do so. Significant amounts of unique variants of each accessory protein having wide variability might contribute significantly to the pathogenicity of SARS-CoV-2. Therefore, our firm conviction that natural weakened stability (if achievable) of SARS-CoV-2 seems to be a far reachable destiny that alarms the danger of the present pandemic scenario due to COVID-19. Also, unique accessory protein variants across individual continents would all be expected to be mixed, while international travels would be restarted without strict protective measures. In this regard, it is our (SACRED, Self-Assembled COVID-19 Research & Education Directive", consisting of international experts in mathematics, physics, computer science, bioinformatics, nanotechnology, structural biology, 245 molecular biology, immunology, and virology) strong recommendation to governmental and non-governmental administrations to take necessary measures to mitigate the spread of COVID-19. SSH conceived the project and carried out the preliminary work. SSH analyzed the results and wrote the primary draft of the article. All authors critically reviewed, edited, and approved the final manuscript. S1 QNE11892 S13 QKO25582 S25 QOH29849 S37 QKG86850 S49 QLB39372 S61 QNO87603 S2 QOI60338 S14 QJS54110 S26 QNA37799 S38 QNA37703 S50 QOF13773 S62 QKV37732 S3 QMX85113 S15 QJC19423 S27 QNR99459 S39 QJA16812 S51 QOH27322 S63 QNO58695 S4 QMU94792 S16 QOI53465 S28 QOE87928 S40 QNM80965 S52 QKQ63440 S64 QNO87963 S5 QIU81889 S17 QJT72174 S29 QMT93272 S41 QOC65745 S53 QKV39531 S65 QNO67179 S6 QNJ45330 S18 QJC21021 S30 QOC65685 S42 QKU31089 S54 QKU30333 S66 QLG76377 S7 QNL36014 S19 QJT72858 S31 QMJ19995 S43 QNU10840 S55 QNN87740 S67 QKV37240 S8 QMU94900 S20 QMI98359 S32 QLI50453 S44 QOF10845 S56 QMS54339 S68 QNO62835 S9 YP 009724394 S21 QOI10363 S33 QOF08397 S45 QOF14025 S57 QMJ00949 S69 QJR87301 S10 QNL98449 S22 QMJ00613 S34 QKV39052 S46 QLC91284 S58 QMT49529 S70 QNV49474 S11 QKO25642 S23 QMU25387 S35 QOJ86810 S47 QOF12309 S59 QNP00779 S71 QNV50338 S12 QKV49390 S24 QNL13170 S36 QMT57060 S48 QKV06503 S60 QJR87841 S72 QMB22615 S1 QKT21007 S27 QJF76096 S53 QNL23941 S79 QKU33322 S105 QOF20590 S131 QNO59248 S2 QMX85114 S28 QOF09958 S54 QOJ40868 S80 QMU25172 S106 QNS28602 S132 QNO66520 S3 QNN90081 S29 QNQ16882 S55 QOF08182 S81 QMS51487 S107 QOF19618 S133 QNO90124 S4 QOI60339 S30 QLH27803 S56 QNA38004 S82 QMU25112 S108 QMS54508 S134 QNO93328 S5 QJZ28119 S31 QOH26975 S57 QOF14542 S83 QNL11851 S109 QIZ14130 S135 QNO64828 S6 QMX85090 S32 QNO32893 S58 QNC69326 S84 QMJ00890 S110 QNM94399 S136 QNO73996 S7 QMU94793 S33 QOF21250 S59 QNL23738 S85 QLG00044 S111 QNS17749 S137 QKV37313 S8 QLH56163 S34 QLJ93776 S60 QOE87249 S86 QMT53402 S112 QLG98244 S138 QNO88516 S9 QLH56283 S35 QOI11612 S61 QOC60864 S87 QOI10196 S113 QNS17689 S139 QJR92882 S10 QLR07201 S36 QMI94146 S62 QMT93717 S88 QNU10973 S114 QNI23442 S140 QNO58696 S11 QLH90095 S37 QJR84962 S63 QNS29326 S89 QNA39599 S115 QMT48978 S141 QNO92404 S12 QNB17764 S38 QMI91915 S64 QOF21190 S90 QOE87321 S116 QMT48966 S142 QLG76138 S13 QNL98462 S39 QNS30322 S65 QNG41678 S91 QNN95802 S117 QOC61296 S143 QNO90292 S14 QJW69144 S40 QNI24809 S66 QNS28494 S92 QMT91425 S118 QMJ01226 S144 QNO58720 S15 QNL35871 S41 QOI60423 S67 QNN95250 S93 QMT91245 S119 QMX86011 S145 QNO64900 S16 YP 009724395 S42 QNT35437 S68 QMU94037 S94 QMU94709 S120 QNM94471 S146 QLG75826 S17 QMS95046 S43 QNI23370 S69 QOE87309 S95 QOE76015 S121 QKS90016 S147 QNO61324 S18 QLF97752 S44 QLC46450 S70 QOJ86811 S96 QOE81298 S122 QIX13980 S148 QNO70240 S19 QKQ30155 S45 QOE44625 S71 QNU10477 S97 QKV41188 S123 QOE81166 S149 QNO80368 S20 QNJ45535 S46 QKU53354 S72 QNQ16954 S98 QOH29130 S124 QLJ58162 S150 QNO66460 S21 QLA46617 S47 QMI90741 S73 QIS61307 S99 QJX74540 S125 QOI60459 S151 QNO96076 S22 QNR60421 S48 QKU33310 S74 QJS54699 S100 QJF76899 S126 QIU81254 S152 QNP02976 S23 QKY65318 S49 QOF07894 S75 QND76351 S101 QMT51066 S127 QNO69016 S153 QNO59080 S24 QJR84386 S50 QMI97185 S76 QOF12514 S102 QNS30406 S128 QNP05940 S154 QNV50135 S25 QMT97821 S51 QNV71003 S77 QNQ17002 S103 QLC48045 S129 QNO69448 S155 QNV50219 S26 QOI53466 S52 QOI10028 S78 QNN86414 S104 QOF08242 S130 QNP07212 S156 QNV49775 S157 QNV49475 S158 QNV49691 QKY59990 S101 QJS54215 S132 QLJ93700 S163 QNG41554 S194 QNO31749 S225 QOH26275 S9 QKS67456 S40 QLQ87577 S71 QLF98084 S102 QKE10935 S133 QOC62516 S164 QLM05764 S195 QNU10705 S226 QOC67026 S10 QLH93429 S41 QKO25747 S72 QLF98048 S103 QJS39497 S134 QMT49694 S165 QNU10765 S196 QNA36464 S227 QNU07492 S11 QLF98036 S42 QKX47995 S73 QLH56099 S104 QNC69819 S135 QKV40164 S166 QNS00146 S197 QLY88564 S228 QIZ16438 S12 QNL35975 S43 QMS50988 S74 QMU94765 S105 QOE86813 S136 QOH26347 S167 QMI95795 S198 QKG90399 S229 QOH27307 S13 QKE61733 S44 QKU37034 S75 QNL36011 S106 QJS53735 S137 QNU11783 S168 QKG64052 S199 QOF18738 S230 QKG86518 S14 QNL35963 S45 QLL26047 S76 QJY40506 S107 QLJ53549 S138 QNL11079 S169 QJW28665 S200 QOF11886 S231 QOH26287 S15 QLH93441 S46 QKJ84956 S77 QMJ01246 S108 QJS54191 S139 QOH26407 S170 QLA47500 S201 QNN96433 S232 QOI09856 S16 QNT09953 S47 QHZ00380 S78 QLH93202 S109 QJW69023 S140 QNL12447 S171 QKN20812 S202 QOH26875 S233 QNL11403 S17 QLF97772 S48 QJD47873 S79 QLH55768 S110 QJS39520 S141 QJV21807 S172 S1 QNN90094 S11 QOI53467 S21 QOJ86812 S31 QNP01153 S2 QOI60340 S12 QNU10478 S22 QMI93943 S32 QLG75935 S3 QKX48960 S13 QNL24002 S23 QOH26412 S33 QNO58697 S4 QNJ45416 S14 QNL11060 S24 QNV70236 S34 QNO90425 S5 QMU84916 S15 QNU10274 S25 QNO30378 S35 QLG75923 S6 YP 009725318 S16 QMT53511 S26 QKE45866 S36 QNO74885 S7 QNN88304 S17 QOI10365 S27 QJC19833 S37 QNV49476 S8 QJQ84777 S18 QOF08027 S28 QKU28444 S9 QNL90926 S19 QMI96613 S29 QJD47604 S10 QKM76816 S20 QKY77886 S30 QKV38827 QNA38594 S100 QMT95795 S125 QOI10114 S150 QOE45090 S175 QNO92610 Figure 11 : Unique variants of accessory protein sequences. 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