Cold-shock induced high-yield protein production in Escherichia coli.
Overexpression of proteins in Escherichia coli at low temperature improves their solubility and stability. Here, we apply the unique features of the cspA gene to develop a series of expression vectors, termed pCold vectors, that drive the high expression of cloned genes upon induction by cold-shock. Several proteins were produced with very high yields, including E. coli EnvZ ATP-binding domain (EnvZ-B) and Xenopus laevis calmodulin (CaM). The pCold vector system can also be used to selectively enrich target proteins with isotopes to study their properties in cell lysates using NMR spectroscopy. We have cloned 38 genes from a range of prokaryotic and eukaryotic organisms into both pCold and pET14 (ref. 3) systems, and found that pCold vectors are highly complementary to the widely used pET vectors.
Qing
Guoliang
G
Department of Biochemistry, Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, New Jersey 08854, USA.
Ma
Li-Chung
LC
Khorchid
Ahmad
A
Swapna
G V T
GV
Mal
Tapas K
TK
Takayama
Masanori Mitta
MM
Xia
Bing
B
Phadtare
Sangita
S
Ke
Haiping
H
Acton
Thomas
T
Montelione
Gaetano T
GT
Ikura
Mitsuhiko
M
Inouye
Masayori
M
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics*; D003001:Cloning, Molecular; D003080:Cold Temperature; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D007553:Isotope Labeling; D008969:Molecular Sequence Data; D019906:Nuclear Magnetic Resonance, Biomolecular; D011401:Promoter Regions, Genetic / Q000235:genetics; D014176:Protein Biosynthesis; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D011506:Proteins; C062260:cold shock protein CS7.4, Bacteria