当前位置： SCI文献检索 > JOURNAL OF MOLECULAR BIOLOGY期刊下所有文献 > The oligomeric structure of vaccinia viral envelope protein A27L is essential for binding to heparin and heparan sulfates on cell surfaces: a structural and functional approach using site-specific mutagenesis.

The oligomeric structure of vaccinia viral envelope protein A27L is essential for binding to heparin and heparan sulfates on cell surfaces: a structural and functional approach using site-specific mutagenesis.

Abstract：

:The soluble domain of the self-assembly vaccinia virus envelope protein A27L, sA27L-aa, consists of a flexible extended coil at the N terminus and a rigid hydrophobic coiled-coil region at the C terminus. In the former, a basic strip of 12 residues is responsible for binding to cell-surface heparan sulfates. Although the latter is believed to mediate self-assembly, its biological role is unclear. However, an in vitro bioassay showed that peptides comprising the 12 residue basic region alone failed to interact with heparin, suggesting that the C-terminal coiled-coil region might serve an indispensable role in biological function. To explore this structural and functional relationship, we performed site-specific mutagenesis in an attempt to specifically disrupt the hydrophobic core of the coiled coil. Three single mutants, L47A, L51A, and L54A, and one triple mutant, L47,51,54A, were expressed and purified from Escherichia coli. The physical properties of the mutants were carefully examined by gel-filtration chromatography, CD, and NMR spectroscopy, and the biological activities were assessed by an in vitro SPR bioassay and three in vivo bioassays: binding to cells, blocking virus infection and blocking cell fusion. We showed that the L47A mutant, which is similar to the parental sA27L-aa in forming a hexamer, is biologically active. L51A and L54A mutants form tetramers and are less active. Notably, in the triple mutant, the self-assembly hydrophobic core structure is uncoiled; as a consequence, the tetrameric structure is biologically inactive. Thus, we conclude that the leucine residues, in particular Leu51 and Leu54, sustain the hydrophobic core structure that is essential for the biological function of vaccinia virus envelope protein A27L, binding to cell-surface heparan sulfate.

journal_name

J Mol Biol

journal_title

Journal of molecular biology

authors

Ho Y,Hsiao JC,Yang MH,Chung CS,Peng YC,Lin TH,Chang W,Tzou DL

doi

10.1016/j.jmb.2005.04.024

keywords:

subject

Has Abstract

pub_date

2005-06-24 00:00:00

pages

1060-71

issue

eissn

0022-2836

issn

1089-8638

pii

S0022-2836(05)00433-X

journal_volume

349

pub_type

杂志文章

在线工具

The oligomeric structure of vaccinia viral envelope protein A27L is essential for binding to heparin and heparan sulfates on cell surfaces: a structural and functional approach using site-specific mutagenesis.