Linker-extended native cyanovirin-N facilitates PEGylation and potently inhibits HIV-1 by targeting the glycan ligand.
Cyanovirin-N (CVN) potently inhibits human immunodeficiency virus type 1 (HIV-1) infection, but both cytotoxicity and immunogenicity have hindered the translation of this protein into a viable therapeutic. A molecular docking analysis suggested that up to 12 residues were involved in the interaction of the reverse parallel CVN dimer with the oligosaccharide targets, among which Leu-1 was the most prominent hot spot residue. This finding provided a possible explanation for the lack of anti-HIV-1 activity observed with N-terminal PEGylated CVN. Therefore, linker-CVN (LCVN) was designed as a CVN derivative with a flexible and hydrophilic linker (Gly4Ser)3 at the N-terminus. The N-terminal α-amine of LCVN was PEGylated to create 10 K PEG-aldehyde (ALD)-LCVN. LCVN and 10 K PEG-ALD-LCVN retained the specificity and affinity of CVN for high mannose N-glycans. Moreover, LCVN exhibited significant anti-HIV-1 activity with attenuated cytotoxicity in the HaCaT keratinocyte cell line and MT-4 T lymphocyte cell lines. 10 K PEG-ALD-LCVN also efficiently inactivated HIV-1 with remarkably decreased cytotoxicity and pronounced cell-to-cell fusion inhibitory activity in vitro. The linker-extended CVN and the mono-PEGylated derivative were determined to be promising candidates for the development of an anti-HIV-1 agent. This derivatization approach provided a model for the PEGylation of biologic candidates without introducing point mutations.
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Jia
J
Chen
School of Pharmaceutical Sciences & Institute of Human Virology, Sun Yat-sen University, Guangzhou, Guangdong, China ; Guangdong Provincial Institutes of Traditional Chinese Medicine, Guangzhou, Guangdong, China.
Dane
D
Huang
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Wei
W
Chen
Department of Molecular Microbiology and Immunology, Nagasaki University, Nagasaki City, Nagasaki Prefecture, Japan.
Chaowan
C
Guo
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Bo
B
Wei
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Chongchao
C
Wu
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Zhou
Z
Peng
Laboratory of Virus Control, Institute for Virus Research, Kyoto University, Kyoto, Japan.
Jun
J
Fan
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Zhibo
Z
Hou
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Yongsheng
Y
Fang
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Yifei
Y
Wang
Department of Molecular Microbiology and Immunology, Nagasaki University, Nagasaki City, Nagasaki Prefecture, Japan.
Kaio
K
Kitazato
Department of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
Guoying
G
Yu
Department of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
Chunbin
C
Zou
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China.
Chuiwen
C
Qian
Institute of Biomedicine & National Engineering Research Center of Genetic Medicine, Jinan University, Guangzhou, Guangdong, China ; Department of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, United States of America.
Sheng
S
Xiong
D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology*; D019281:Dimerization; D015195:Drug Design; D015658:HIV Infections / Q000188:drug therapy*; D015497:HIV-1 / Q000187:drug effects*; D006801:Humans; D008958:Models, Molecular; D062105:Molecular Docking Simulation; D009844:Oligosaccharides / Q000378:metabolism*; D011092:Polyethylene Glycols / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods
D001426:Bacterial Proteins; D002352:Carrier Proteins; D009844:Oligosaccharides; C107733:cyanovirin N; D011092:Polyethylene Glycols