Crystallographic and Glycan Microarray Analysis of Human Polyomavirus 9 VP1 Identifies N -Glycolyl Neuraminic Acid as a Receptor Candidate

Abstract

Human polyomavirus 9 (HPyV9) is a closely related homologue of simian B-lymphotropic polyomavirus (LPyV). In order to define the architecture and receptor binding properties of HPyV9, we solved high-resolution crystal structures of its major capsid protein, VP1, in complex with three putative oligosaccharide receptors identified by glycan microarray screening. Comparison of the properties of HPyV9 VP1 with the known structure and glycan-binding properties of LPyV VP1 revealed that both viruses engage short sialylated oligosaccharides, but small yet important differences in specificity were detected. Surprisingly, HPyV9 VP1 preferentially binds sialyllactosamine compounds terminating in 5-N-glycolyl neuraminic acid (Neu5Gc) over those terminating in 5-N-acetyl neuraminic acid (Neu5Ac), whereas LPyV does not exhibit such a preference. The structural analysis demonstrated that HPyV9 makes specific contacts, via hydrogen bonds, with the extra hydroxyl group present in Neu5Gc. An equivalent hydrogen bond cannot be formed by LPyV VP1. IMPORTANCEThe most common sialic acid in humans is 5-N-acetyl neuraminic acid (Neu5Ac), but various modifications give rise to more than 50 different sialic acid variants that decorate the cell surface. Unlike most mammals, humans cannot synthesize the sialic acid variant 5-N-glycolyl neuraminic acid (Neu5Gc) due to a gene defect. Humans can, however, still acquire this compound from dietary sources. The role of Neu5Gc in receptor engagement and in defining viral tropism is only beginning to emerge, and structural analyses defining the differences in specificity for Neu5Ac and Neu5Gc are still rare. Using glycan microarray screening and high-resolution protein crystallography, we have examined the receptor specificity of a recently discovered human polyomavirus, HPyV9, and compared it to that of the closely related simian polyomavirus LPyV. Our study highlights critical differences in the specificities of both viruses, contributing to an enhanced understanding of the principles that underlie pathogen selectivity for modified sialic acids.

Structure Analysis of the Major Capsid Proteins of Human Polyomaviruses 6 and 7 Reveals an Obstructed Sialic Acid Binding Site

“…1C). Typically, this platform binds glycan receptors that terminate in sialic acid (Neu5Ac), but sequences and linkages of the recognized oligosaccharides differ among polyomaviruses, leading to specific interactions with a small subset of sialylated glycans in each case (28)(29)(30)(32)(33)(34)(35)(36). However, despite these differences, the location of the sialic acid binding site is conserved among the polyomaviruses for which VP1 structures have been available to date.…”

“…Taking these data together, the elongated HI-loops and the truncated BC2-loops of HPyV6 (and also HPyV7) lead to a profoundly altered surface loop network. The HI-loop participates in the recognition of sialylated glycan receptors in all sialic-acid binding polyomaviruses whose structures have been determined to date (28)(29)(30)(32)(33)(34)(35), by contributing parts of the shallow receptor binding groove on the protein The loops are colored as described for panels A and B, respectively. VP1 monomer structures were superposed using the secondary-structure-matching (SSM) superposition tool (45) in the program Coot (42).…”

“…However, despite these differences, the location of the sialic acid binding site is conserved among the polyomaviruses for which VP1 structures have been available to date. The sialic acid binding site is typically located in a recessed area at the junction of the BC1-, BC2-, DE-, and HI-loops of a VP1 monomer, and additional contacts are contributed by the BC2-and DE-loops of counterclockwise (ccw) and clockwise (cw) neighboring monomers, respectively (28)(29)(30)(32)(33)(34)(35). In order to assess the ability of the HPyV6 and HPyV7 surface loops to form such a sialic acid binding site, we compared the conformations and lengths of their Crystal Structures of HPyV6 and HPyV7 VP1 Pentamers surface-exposed loops with the equivalent loops in WUPyV and KIPyV, as well as in MCPyV and SV40.…”

“…For each virus, they form a unique virus-host interaction platform that determines host range, cell tropism, viral spread, and pathogenicity. Engagement of sialylated glycan motifs during cell attachment and entry is a common feature of the better-studied orthopolyomaviruses (28)(29)(30)(32)(33)(34)(35)(36). In contrast, the routes of infection, transmission, cell tropism, receptor specificity, and entry pathways of wukipolyomaviruses remain largely mysterious.…”

“…The presence and roles of VP2 and VP3 seem to differ among polyomavirus species (27). All known structures of polyomavirus VP1 show extended and structurally variable surface loops that emanate from a conserved ␤-sheet core structure formed by strands B, I, D, and G and strands C, H, E, and F (25,26,(28)(29)(30)(31)(32)(33)(34)(35). These surface loops, named BC-, DE-, HI-, and EF-loops after the ␤-strands connected by them, are chiefly responsible for viral antigenicity.…”