Taiwan cobra chymotrypsin inhibitor: cloning, functional expression and gene organization.
A cDNA encoding chymotrypsin inhibitor was constructed from the cellular RNA isolated from the venom glands of Naja atra (Taiwan cobra). The resultant amino acid sequence showed that the mature protein is comprised of 57 amino acid residues with six cysteine residues. Cloned protein was expressed and isolated from the inclusion bodies of E. coli and refolded into a functional protein in vitro. Deleting the first three residues at its N-terminus caused a moderate increase in the inhibitory constant (K(i)) against chymotrypsin. The genomic DNA encoding the chymotrypsin inhibitor was amplified by PCR. The gene shares virtually an identical structural organization with the beta-bungarotoxin B1 chain (a snake Kunitz/BPTI neurotoxic homolog) gene. Moreover, the overall sequence identity of the N. atra chymotrypsin inhibitor and beta-bungarotoxin B1 chain genes was up to 83%. These findings strongly suggest that snake Kunitz/BPTI protease inhibitors and neurotoxic homologs may have originated from a common ancestor.
Cheng
Yun-Ching
YC
Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung 804, Taiwan.
Yan
Fang-Jiun
FJ
Chang
Long-Sen
LS
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002038:Bungarotoxins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002918:Chymotrypsin / Q000037:antagonists & inhibitors* / Q000737:chemistry; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics / Q000302:isolation & purification; D004546:Elapid Venoms / Q000737:chemistry*; D017815:Elapidae / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D008969:Molecular Sequence Data; D016296:Mutagenesis; D016133:Polymerase Chain Reaction; D011480:Protease Inhibitors / Q000378:metabolism*; D013624:Taiwan
D002038:Bungarotoxins; D018076:DNA, Complementary; D004546:Elapid Venoms; D011480:Protease Inhibitors; D002918:Chymotrypsin