Construction of Isopeptide Bridge-tethered NHR-trimeric Coiled-coil in MERS-CoV Membrane Fusion Protein†

doi:10.7503/cjcu20160237

Abstract

Abstract:

In Middle East respiratory syndrome coronavirus(MERS-CoV) infection, the formation of a coiled-coil six-helical bundle(6-HB) between three C-terminal heptad repeats(CHRs) and an N-terminal heptad repeat(NHR) trimer of the MERS-CoV spike protein S2 subunit provided the energy necessary for virus-cell membrane fusion. Mimicry of the NHR-helical trimers in the MERS-CoV membrane fusion protein for the discovery of antiviral therapeutics hampered because of the strong aggregation properties of synthetic NHR-based peptides taken out of their parent protein structure. Herein, the article presents an efficient strategy to recapi-tulate MERS-CoV NHR α-helical trimers via combining the concepts of grafting the NHR-derived peptides onto an exogenous trimerization motif with stabilization of the trimeric oligomers through isopeptide bonds. The covalent bridge was introduced by an acyl transfer reaction between lysine and glutamic acid in specific amino acid sites with thio-easter modified. The designed isopeptide bridge-stabilized chimeric trimers has strong potential for the development potent MERS-CoV fusion inhibitors.

Key words: Middle East respiratory syndrome coronavirus, Fusion protein, Peptide, Isopeptide bond

CLC Number:

O629.72

TrendMD:

ZHENG Xi, LIANG Guodong, WANG Chao, LIU Keliang. Construction of Isopeptide Bridge-tethered NHR-trimeric Coiled-coil in MERS-CoV Membrane Fusion Protein^†[J]. Chem. J. Chinese Universities, 2016, 37(9): 1643.

Figures/Tables 4

Scheme 1 Structure of 6-helix bundle(6-HB) (A) Side view of MERS-CoV S2 subunit 6-HB structure(PDB: 4mod), peripheral helix present the C-terminal heptad repeat(CHR) in MERS-CoV, while the central helix trimer are N-terminal heptad repeat(NHR); (B) top view of 6-HB show the amino acid in a, d site(cyan) and e, g site.

Scheme 2 N-helix trimer stabled by isopeptide (A) Isopeptide bond was formed between lysine in e site and glutamic acid in g' site of adjacent peptide chain; (B) schematic representation of isopeptide bond formation via an interhelical acyl transfer reaction.

Fig.1 RP-HPLC traces(A—C) and mass spectrometry(D—I) of isopeptide bond forming reaction RP-HPLC traces for the acyl transfer reaction of (IZN21L)3(A), (IZN21M)3(B) and (IZN21R)3(C) from t=2.5 to 150 h. Absorbance was monitored at 210 nm. (A—C): a. The reaction material; b. the reaction product; c. by-products. Mass spectrometry of monomer peptide, the reaction materal of (IZN21L)3(D), (IZN21M)3(E), (IZN21R)3(F) and reaction products, the trimeric peptide (IZN21L)3(G), (IZN21M)3(H) and (IZN21R)3(I).

Table 1 Designed peptide sequences derived from HR1P

Name	Sequence^*
	e f g a b c de f g
HR1P(998—1039)	ANK FNQALGA MQTGFTTTNEAFQKVQDAVNNNAQALSKLASE
(IZN21L)₃	(IZm---ANK FNQALGA MQTGFTTTNEA)₃
(IZN21M)₃	(IZm-----------------LGA MQTGFTTTNEAFQKVQDA)₃
(IZN21R)₃	(IZm-------------------------------------------FQKVQDAVNNNAQALSKLASE)₃

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