[Biological synthesis and purification of spider dragline silk protein polymers containing RGD three peptide].
Spider dragline silk is one of most perfect fibrous proteins in nature. As biomaterials, it has a wide application in tissue engineering due to its unique mechanical properties, good biocompatibility, slow degradation. In this paper, based on the highly repetitive sequence of spider dragline silk and with the introduced RGD peptide codons which involve cell adhesion, the DNA monomer sequence encoding RGD-spider dragline silk was synthesized, and then was used to construct the multimers by the strategy of "head to tail"; the multimers were ligated into prokaryotic expression vector pET-30a, and then the B121 (DE3) pLyS, were transformed the expression of recombinant protein was induced by the addition of IPTG. SDS-PAGE analysis shows that the molecular weight of products expressed here are 35KD and 60KD respectively in agreement with the desired. Western assay was used for determining the specification of products. Further, the purification process was groped for the producing of large quantity of synthetic proteins through high density fermentation.
Li
Min
M
College of Bioengineering, Fujian Normal University, Fuzhou 350007, China. mli@fjnu.edu.cn
Huang
Jiankun
J
Tu
Guiyun
G
Huang
Xi
X
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001672:Biocompatible Materials; D003001:Cloning, Molecular; D005349:Fibroins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification*; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000096:biosynthesis* / Q000302:isolation & purification*; D011108:Polymers / Q000302:isolation & purification; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D013112:Spiders
D001672:Biocompatible Materials; D009842:Oligopeptides; D011108:Polymers; D011994:Recombinant Proteins; C076815:spidroin 2; C047981:arginyl-glycyl-aspartic acid; D005349:Fibroins