Switch to
| Predicate | Object |
|---|---|
| rdf:type | |
| lifeskim:mentions | |
| pubmed:issue |
14
|
| pubmed:dateCreated |
1995-5-15
|
| pubmed:abstractText |
The deduced primary sequence of the cytoplasmic protein-tyrosine kinase domain of the insulin receptor contains a conserved kinase homology region (receptor residues 1002-1257) flanked by a juxtamembrane region and a C-terminal tail. A soluble 48-kDa derivative (residues 959-1355) containing these regions but lacking the first six residues of the juxtamembrane region had earlier been synthesized in Sf9 cells using a baculovirus expression system. The catalytic core of the kinase domain was studied first by proteolytic analysis of the 48-kDa kinase and then by expressing a series of truncated kinase domains in transiently transfected COS cells. Based on these studies, two core kinases of 34 (residues 985-1283) and 35 (residues 978-1283) kDa, respectively, were overexpressed in Sf9 cells. Biochemical characterization of the 35-kDa kinase revealed that the core kinase conserved the major functional properties of the native receptor kinase domain. Activity of the 35-kDa kinase toward a synthetic peptide increased more than 200-fold upon autophosphorylation, which occurred exclusively at Tyr-1158, Tyr-1162, and Tyr-1163; the largest increase was observed between bis- and trisphosphorylation of the kinase. The activated 35- and 48-kDa kinases were similar with respect to specific activity and ATP and Mg2+ requirements for peptide phosphorylation. Moreover, autophosphorylation appeared to initiate predominantly at Tyr-1162, immediately followed by phosphorylation at Tyr-1158 and then at Tyr-1163. The rate of autophosphorylation was dependent on enzyme concentration, consistent with a trans-phosphorylation mechanism. Finally, the 35-kDa kinase was crystallized, making possible elucidation of its three-dimensional structure by x-ray crystallography.
|
| pubmed:language |
eng
|
| pubmed:journal | |
| pubmed:citationSubset |
IM
|
| pubmed:chemical | |
| pubmed:status |
MEDLINE
|
| pubmed:month |
Apr
|
| pubmed:issn |
0021-9258
|
| pubmed:author | |
| pubmed:issnType |
Print
|
| pubmed:day |
7
|
| pubmed:volume |
270
|
| pubmed:owner |
NLM
|
| pubmed:authorsComplete |
Y
|
| pubmed:pagination |
8122-30
|
| pubmed:dateRevised |
2009-11-19
|
| pubmed:meshHeading |
pubmed-meshheading:7713916-Amino Acid Sequence,
pubmed-meshheading:7713916-Animals,
pubmed-meshheading:7713916-Baculoviridae,
pubmed-meshheading:7713916-Base Sequence,
pubmed-meshheading:7713916-Catalysis,
pubmed-meshheading:7713916-Cell Line,
pubmed-meshheading:7713916-Cloning, Molecular,
pubmed-meshheading:7713916-Crystallization,
pubmed-meshheading:7713916-DNA, Complementary,
pubmed-meshheading:7713916-Humans,
pubmed-meshheading:7713916-Molecular Sequence Data,
pubmed-meshheading:7713916-Phosphorylation,
pubmed-meshheading:7713916-Receptor, Insulin,
pubmed-meshheading:7713916-Spodoptera,
pubmed-meshheading:7713916-Tyrosine
|
| pubmed:year |
1995
|
| pubmed:articleTitle |
Expression, characterization, and crystallization of the catalytic core of the human insulin receptor protein-tyrosine kinase domain.
|
| pubmed:affiliation |
W. M. Keck Center for Genome Informatics, Texas A & M University, Houston 77030, USA.
|
| pubmed:publicationType |
Journal Article,
Research Support, U.S. Gov't, P.H.S.,
Research Support, U.S. Gov't, Non-P.H.S.,
Research Support, Non-U.S. Gov't
|