Purification and cDNA cloning of a four-domain Kazal proteinase inhibitor from crayfish blood cells.
A cDNA with an open reading frame of 684 base pairs was isolated from a library from blood cells of the crayfish Pacifastacus leniusculus. It codes for a signal sequence and a mature protein of 209 amino acids with a predicted molecular mass of 22.7 kDa. The amino acid sequence consists of four repeated stretches (45-73% identical to each other), indicating that the protein has four domains. The domains have significant sequence similarity to serine proteinase inhibitors of the Kazal family. The three first domains have a leucine residue in the putative reactive site, suggesting that the protein is a chymotrypsin inhibitor. A monomeric 23-kDa proteinase inhibitor, which by amino terminal sequencing of the mature protein was confirmed to be the cloned Kazal inhibitor, was purified from crayfish blood cells. It inhibited chymotrypsin or subtilisin, but not trypsin, elastase or thrombin. The inhibitor seemed to form a 1:1 complex with chymotrypsin or subtilisin. This protein seems to be the first described Kazal inhibitor from blood cells of any animal and the first one with four domains.
Johansson
M W
MW
Department of Physiological Botany, University of Uppsala, Sweden.
Keyser
P
P
Söderhäll
K
K
D000595:Amino Acid Sequence; D000818:Animals; D003400:Astacoidea / Q000737:chemistry*; D001483:Base Sequence; D001773:Blood Cells / Q000737:chemistry; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics*; D004591:Electrophoresis, Polyacrylamide Gel; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016415:Sequence Alignment; D014359:Trypsin Inhibitor, Kazal Pancreatic / Q000097:blood / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification
D018076:DNA, Complementary; D014359:Trypsin Inhibitor, Kazal Pancreatic