Expression, purification, and bioactivity of GST-fused v-Src from a bacterial expression system.
v-Src is a non-receptor protein tyrosine kinase involved in many signal transduction pathways and closely related to the activation and development of cancers. We present here the expression, purification, and bioactivity of a GST (glutathione S-transferase)-fused v-Src from a bacterial expression system. Different culture conditions were examined in an isopropyl beta-D-thiogalactopyranoside (IPTG)-regulated expression, and the fused protein was purified using GSH (glutathione) affinity chromatography. ELISA (enzyme-linked immunosorbent assay) was employed to determine the phosphorylation kinase activity of the GST-fused v-Src. This strategy seems to be more promising than the insect cell system or other eukaryotic systems employed in earlier Src expression.
Gong
Xing-Guo
XG
Institute of Biomacromolecule and Enzyme Engineering, School of Life Sciences, Zhejiang University, Hangzhou 310027, China. gongxg@cls.zju.edu.cn
Ji
Jing
J
Xie
Jie
J
Zhou
Yuan
Y
Zhang
Jun-Yan
JY
Zhong
Wen-Tao
WT
D001426:Bacterial Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D005982:Glutathione Transferase / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D015688:Oncogene Protein pp60(v-src) / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D015202:Protein Engineering / Q000379:methods*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase; D015688:Oncogene Protein pp60(v-src)