Molecular evolution of cystine-stabilized miniproteins as stable proteinaceous binders.
Small cystine-stabilized proteins are desirable scaffolds for therapeutics and diagnostics. Specific folding and binding properties of the proteinaceous binders can be engineered with combinatorial protein libraries in connection with artificial molecular evolution. The combinatorial protein libraries are composed of scaffold variants with random sequence variation, which inevitably produces a portion of the library sequences incompatible with the parent structure. Here, we used artificial molecular evolution to elucidate structure-determining residues in a smallest cystine-stabilized scaffold. The structural determinant information was then applied to designing cystine-stabilized miniproteins binding to human vascular endothelial growth factor. This work demonstrated a general methodology on engineering artificial cystine-stabilized proteins as antibody mimetics with simultaneously enhanced folding and binding properties.
Chang
Hung-Ju
HJ
Genomics Research Center, Academia Sinica, Taipei 115, Taiwan.
Hsu
Hung-Ju
HJ
Chang
Chi-Fon
CF
Peng
Hung-Pin
HP
Sun
Yi-Kun
YK
Yu
Hui-Ming
HM
Shih
Hsi-Chang
HC
Song
Chun-Ying
CY
Lin
Yi-Ting
YT
Chen
Chu-Chun
CC
Wang
Chia-Hung
CH
Yang
An-Suei
AS
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D003553:Cystine / Q000737:chemistry*; D004220:Disulfides / Q000737:chemistry; D019143:Evolution, Molecular; D006801:Humans; D008969:Molecular Sequence Data; D019151:Peptide Library; D011485:Protein Binding / Q000235:genetics; D011487:Protein Conformation; D015202:Protein Engineering / Q000379:methods*; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism
D004220:Disulfides; D019151:Peptide Library; D011506:Proteins; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D003553:Cystine