Insights into autoregulation from the crystal structure of twitchin kinase.
Many protein kinases are self-regulated by an intrasteric mechanism where part of the enzyme's structure directly inhibits the active site. This inhibitory structure is called a pseudosubstrate and specific regulators are required to remove it from the active site to allow substrates access. Removal of the pseudosubstrate sequence from members of the myosin light-chain kinase subfamily, including twitchin kinase, activates them but it is not known whether the pseudosubstrate sequence binds to the active site. Native twitchin is a 753K protein (6,839 residues) located in muscle A-bands of the nematode Caenorhabditis elegans and because of its size has not been easy to study. We have determined the crystal structure, refined to 2.8 A resolution, of a recombinant fragment (residues 5,890 to 6,262) of twitchin kinase that contains the catalytic core and a 60 residue carboxy-terminal tail. The C-terminal tail extends through the active site, wedged between the small and large lobes of the structure and making extensive contacts with the catalytic core which accounts for autoinhibition and provides direct support for the intrasteric mechanism of protein kinase regulation.
Hu
S H
SH
St. Vincent's Institute of Medical Research, Fitzroy, Victoria, Australia.
Parker
M W
MW
Lei
J Y
JY
Wilce
M C
MC
Benian
G M
GM
Kemp
B E
BE
D000595:Amino Acid Sequence; D000818:Animals; D017173:Caenorhabditis elegans / Q000201:enzymology; D029742:Caenorhabditis elegans Proteins; D002148:Calmodulin-Binding Proteins; D003196:Computer Graphics; D018360:Crystallography, X-Ray; D017868:Cyclic AMP-Dependent Protein Kinases / Q000737:chemistry; D015801:Helminth Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry; D011487:Protein Conformation; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry
D029742:Caenorhabditis elegans Proteins; D002148:Calmodulin-Binding Proteins; D015801:Helminth Proteins; D009124:Muscle Proteins; D010446:Peptide Fragments; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; C489233:unc-22 protein, C elegans; D011494:Protein Kinases; D017868:Cyclic AMP-Dependent Protein Kinases