乙型肝炎病毒(HBV)表达共末端大(L),中间(M),和含有preS1/preS2/S的小(S)包膜蛋白,preS2/S,和S域单独,分别。S和preS1结构域介导与硫酸乙酰肝素蛋白聚糖和牛磺胆酸钠共转运多肽(NTCP)的序列病毒体附着,分别,可被抗S和抗preS1抗体阻断。抗preS2抗体如何中和HBV感染性仍然是神秘的。慢性HBV感染的晚期通常选择突变的preS2翻译起始密码子,以防止M蛋白表达,或框内preS2缺失以缩短L和M蛋白。当引入基因型C或D的感染性克隆时,M-负突变和大多数5'preS2缺失都维持了病毒体的产生。这种突变子代病毒颗粒在NTCP重建的HepG2细胞中具有感染性。对基因型D克隆进行中和实验。尽管仍然易感抗preS1和抗S中和抗体,M-负突变体仅被测试的两种抗preS2抗体部分中和,而preS2缺失突变体具有抗性。通过感染实验,使用具有丢失与增加的M蛋白表达的病毒颗粒,或仅存在于L或M蛋白上的中和逃逸preS2缺失,我们发现全长L和M蛋白均有助于两种抗preS2抗体中和病毒.因此,免疫逃逸可能是选择M-负突变的驱动力,尤其是preS2删除。L和M蛋白均可介导抗preS2抗体的中和,这一事实可能揭示了潜在的分子机制。重要的大(L),中间(M),和乙型肝炎病毒(HBV)的小(S)包膜蛋白含有preS1/preS2/S,preS2/S,和S域单独,分别。硫酸乙酰肝素蛋白聚糖和牛磺胆酸钠协同转运多肽(NTCP)作为低和高亲和力HBV受体的发现可以解释抗S和抗preS1抗体的中和潜力,分别,但是抗preS2中和抗体是如何工作的仍然是神秘的。在这项研究中,我们在基因型D的背景下发现了两个M-负突变体,在NTCP重建的HepG2细胞中部分逃脱了两个抗preS2中和抗体,而几个天然存在的preS2缺失突变体逃脱了这两种抗体。通过点突变来消除或增强M蛋白的表达,通过选择性地将preS2缺失引入L或M蛋白,我们发现抗preS2抗体与L和M蛋白的结合有助于中和野生型HBV感染性。我们的发现可能揭示了抗preS2抗体中和HBV感染性的可能机制。
Hepatitis B virus (HBV) expresses co-terminal large (L), middle (M), and small (S) envelope proteins containing preS1/preS2/S, preS2/S, and S domain alone, respectively. S and preS1 domains mediate sequential virion attachment to heparan sulfate proteoglycans and sodium taurocholate cotransporting polypeptide (NTCP), respectively, which can be blocked by anti-S and anti-preS1 antibodies. How anti-preS2 antibodies neutralize HBV infectivity remains enigmatic. The late stage of chronic HBV infection often selects for mutated preS2 translation initiation codon to prevent M protein expression, or in-frame preS2 deletions to shorten both L and M proteins. When introduced to infectious clone of genotype C or D, both M-minus mutations and most 5\' preS2 deletions sustained virion production. Such mutant progeny viral particles were infectious in NTCP-reconstituted HepG2 cells. Neutralization experiments were performed on the genotype D clone. Although remaining susceptible to anti-preS1 and anti-S neutralizing antibodies, M-minus mutants were only partially neutralized by two anti-preS2 antibodies tested while preS2 deletion mutants were resistant. By infection experiments using viral particles with lost versus increased M protein expression, or a neutralization escaping preS2 deletion only present on L or M protein, we found that both full-length L and M proteins contributed to virus neutralization by the two anti-preS2 antibodies. Thus, immune escape could be a driving force for the selection of M-minus mutations, and especially preS2 deletions. The fact that both L and M proteins could mediate neutralization by anti-preS2 antibodies may shed light on the underlying molecular mechanism.IMPORTANCEThe large (L), middle (M), and small (S) envelope proteins of hepatitis B virus (HBV) contain preS1/preS2/S, preS2/S, and S domain alone, respectively. The discovery of heparan sulfate proteoglycans and sodium taurocholate cotransporting polypeptide (NTCP) as the low- and high-affinity HBV receptors could explain neutralizing potential of anti-S and anti-preS1 antibodies, respectively, but how anti-preS2 neutralizing antibodies work remains enigmatic. In this study, we found two M-minus mutants in the context of genotype D partially escaped two anti-preS2 neutralizing antibodies in NTCP-reconstituted HepG2 cells, while several naturally occurring preS2 deletion mutants escaped both antibodies. By point mutations to eliminate or enhance M protein expression, and by introducing preS2 deletion selectively to L or M protein, we found binding of anti-preS2 antibodies to both L and M proteins contributed to neutralization of wild-type HBV infectivity. Our finding may shed light on the possible mechanism(s) whereby anti-preS2 antibodies neutralize HBV infectivity.