[Studies on expression, purification, crystal growth and optimization of putative transcription factor LytR from Streptococcus pneumoniae].
The aim of the present study was to obtain the crystal of transcription factor LytR of streptococcus pneumoniae for X-ray crystal structure and function analysis. The LytR gene of D39 strains from Streptococcus pneumoniae (S. pn) was cloned into the prokaryotic expression vector pET32a(+), then overexpression was obtained in the E. coli BL21 (DE3) through transformation of the recombinant plasmid that had been verified by colony PCR and sequencing. Soluble fusion protein with His-tag highly expressed by the induction of 0.5 mmol/L IPTG and was purified by a three step procedure, the purity of the purified LytR recombinant protein was over 90%. Preliminary screening of crystallization conditions was performed using the hanging-drop vapour-diffusing method with Hampton Crystal screen and PEG screen kits. The protein crystals X-ray diffraction data were collected from a single crystal and more stick crystals whose X-ray diffraction reached 4.0 A were obtained. These works laid the foundation for further research on the 3D structure of putative transcription factor LytR and its biological aspects.
Department of Medicine Laboratory, Affliated Hospital of Zunyi Medical College, Zunyi 563003, China.
Xun
X
Min
Wen
W
Zhong
Shasha
S
Zhao
Jie
J
Dong
Shanshan
S
Dong
Aie
A
Zhou
Wenjuan
W
Yan
Deqiang
D
Wang
D001426:Bacterial Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D013296:Streptococcus pneumoniae / Q000235:genetics* / Q000378:metabolism; D014157:Transcription Factors / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification
D001426:Bacterial Proteins; C500439:CpsA protein, Streptococcus pneumoniae; D011994:Recombinant Proteins; D014157:Transcription Factors