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rdf:type |
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lifeskim:mentions |
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pubmed:issue |
3
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pubmed:dateCreated |
1993-7-27
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pubmed:abstractText |
The gene encoding for a pumpkin (Curcubita maxima) trypsin inhibitor CMTI-V was synthesized chemically. The synthetic gene was prepared from four overlapping oligonucleotides by overlapping extension. The synthetic gene was amplified by polymerase chain reaction, cloned into a T7 expression vector and expressed in Escherichia coli as a fusion protein. The clone, namely 70-1, encoded a fusion protein containing 7 amino acid residues of the N-terminus of the bacterial protein rho 10 and the entire 68 residues of CMTI-V. The wild-type fusion protein constituted approximately 15% of the total bacterial protein mass and was purified to homogeneity in a single step by antibody affinity chromatography. The wild-type fusion protein possesses inhibitory activity toward trypsin and beta-Factor XIIa, but to a lesser extent when compared to the natural CMTI-V. A mutant, T43A, in which threonine at position 43 (P2 position) was replaced by alanine, was constructed. This mutant showed considerably lower specific inhibitory activity toward both trypsin and beta-Factor XIIa.
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pubmed:language |
eng
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pubmed:journal |
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pubmed:citationSubset |
IM
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pubmed:chemical |
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pubmed:status |
MEDLINE
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pubmed:month |
Jun
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pubmed:issn |
1046-5928
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pubmed:author |
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pubmed:issnType |
Print
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pubmed:volume |
4
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pubmed:owner |
NLM
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pubmed:authorsComplete |
Y
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pubmed:pagination |
215-22
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pubmed:dateRevised |
2006-11-15
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pubmed:meshHeading |
pubmed-meshheading:8518561-Amino Acid Sequence,
pubmed-meshheading:8518561-Base Sequence,
pubmed-meshheading:8518561-Cloning, Molecular,
pubmed-meshheading:8518561-Dose-Response Relationship, Drug,
pubmed-meshheading:8518561-Escherichia coli,
pubmed-meshheading:8518561-Factor XIIa,
pubmed-meshheading:8518561-Frameshift Mutation,
pubmed-meshheading:8518561-Genes, Plant,
pubmed-meshheading:8518561-Genes, Synthetic,
pubmed-meshheading:8518561-Genetic Vectors,
pubmed-meshheading:8518561-Molecular Sequence Data,
pubmed-meshheading:8518561-Mutagenesis, Site-Directed,
pubmed-meshheading:8518561-Plant Proteins,
pubmed-meshheading:8518561-Recombinant Proteins,
pubmed-meshheading:8518561-Sequence Homology, Amino Acid,
pubmed-meshheading:8518561-Solubility,
pubmed-meshheading:8518561-Trypsin,
pubmed-meshheading:8518561-Trypsin Inhibitors
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pubmed:year |
1993
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pubmed:articleTitle |
Chemical synthesis, molecular cloning, overexpression, and site-directed mutagenesis of the gene coding for pumpkin (Curcubita maxima) trypsin inhibitor CMTI-V.
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pubmed:affiliation |
Department of Chemistry, Western Illinois University, Macomb 61455.
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pubmed:publicationType |
Journal Article,
Comparative Study,
Research Support, Non-U.S. Gov't
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