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  • Previously the N glycan in V of HXB


    Previously, the N-glycan in V3 of HXB2, HXB10, LAI and BRU strains was reported to be dispensable for their infectivity to T cell lines, although detailed growth kinetics and experimental conditions were not described 4, 5, 6, 7, 8. The experiments appear to have been done by infection initiated with only a single virus dose that was not specified clearly. We have to note that differences between the wild-type and mutant viruses were not observed with higher input virus doses, suggesting that the effect of glycan removal would essentially be small in a single step replication, and become appreciable only after numerous rounds of replication. Even though small to a single replication cycle, the contribution of the glycan would be great enough to in vivo pathogenesis which involves persistence of highly active viral replications for a long period. It was previously noted that the N-glycosylation site in V3 is less conserved among T-tropic isolates than among M-tropic isolates [5]. This is apparently paradoxical if the N-glycan is important for CXCR-4- but not CCR-5-dependent entry as proposed in this study. However, inspection of V3 sequences revealed that the loss of the N-glycan is closely associated with a basic amino Omecamtiv mecarbil where substitution at the 11th position. Furthermore, the basic amino acid substitution at the 11th position in NL43HNT partially but significantly restored the infectious capacity. Therefore, the N-glycan in V3 seems to be dispensable only in the particular context of the peptide backbone, and plays an important role for CXCR-4-dependent membrane fusion when the 11th position of V3 is occupied by a non-basic residue. On the other hand, removal of the N-glycan from the V3 loop was reported to increase V3-specific antigenicity 5, 6, 7, 8. We could confirm this since the NL43HNT showed increased sensitivity to the V3-specific neutralizing monoclonal antibody 902 (data not shown). Back et al. [5]further raised the view that the N-glycan protects the V3 loop from attack by antibodies and hence is dispensable in patients with compromised immunity. However, in view of our present data showing a functional requirement of the glycan, the situation does not appear to be that simple. Their view may be applicable only to M-tropic strains whose N-glycan in V3 does not appear to be functionally important. The V3 loop was suggested to be involved in the gp120-co-receptor interaction 40, 41. Our present study demonstrated that there is a strain-specific requirement of the V3 glycan for this interaction. It is interesting but difficult to explain why and how the V3 glycan contributes to the infection process dependent on CXCR-4 but not CCR-5. It was previously reported that a subtle charge change in V3 of the T-tropic HIV-1 strain SF2 caused a drastic reduction of viral replicating capability in T cell lines, whereas the corresponding charge change in the M-tropic strain SF162 did not affect macrophage tropism [18]. It is thus possible that the V3 of T-tropic strains should be structurally more stringent for co-receptor recognition than that of M-tropic strains, and therefore the removal of N-glycan from the former could make an impact on virus-co-receptor interaction and infectivity. The N-glycan on V3 is supposed to be a complex type and thus negatively charged [1]. A single N-linked sugar chain possesses a molecular mass of 2000–3000. These physical and structural properties may somehow be required for V3 to fully interact with CXCR-4. However, the sugar chain may not be directly involved in the interaction, since a basic amino acid substitution at a nearby site partially compensated for the loss of the glycan. It is puzzling if the presence of a negative charge (in the glycan) and a basic amino acid substitution for a neutral nearby residue are both factors leading to upregulation of CXCR-4-dependent fusion. Increase of hydrophilicity is a common feature shared by both. V3 glycosylation and the basic amino acid substitution may thus facilitate V3 exposure for better interaction with CXCR-4. In any case, a basic amino acid substitution at the 11th position appears to be an apt change in vivo for this particular N-glycan-negative virus to acquire a sufficiently high infectivity or for the virus to evade clearance due to infectivity reduction caused by spontaneous removal of the glycan.