Molecular cloning and expression of rabbit heparin cofactor II: a plasma thrombin inhibitor highly conserved between species.
Heparin cofactor II (HCII), a circulating plasma protein that inhibits thrombin, is a member of the serine proteinase (serpin) family of proteins. The extent to which HCII structure is conserved across species lines was investigated, by obtaining cDNA clones encoding rabbit HCII. Overlapping clones corresponding to rabbit HCII were obtained by the combined use of hybridization screening of a rabbit liver cDNA library, and by rapid amplification of cDNA ends (RACE). The consensus sequence obtained spans 2178 nucleotides, and is comprised of a 5' untranslated region of 77 nucleotides, an open reading frame of 1440 nucleotides, and a 3' untranslated region of 661 nucleotides that concludes with a poly A tract. The open reading frame is subdivided into a secretory signal sequence of 19 amino acids, and a mature protein of 461 amino acids. Within the region comprising the mature protein, 87% of the amino acid residues are identical to those seen in human HCII. Expression of an appropriately modified form of the rabbit HCII clone in an in vitro reticulocyte expression system yielded two major polypeptides, of 60 and 56 kD respectively, both of which were able to form SDS-stable complexes with human alpha-thrombin, in a reaction accelerated by dermatan sulphate. The remarkable degree of homology observed between rabbit HCII and its human counterpart, indicating a high degree of conservation of structure through evolution, suggests an important function of HCII in hemostasis.
Canadian Red Cross Society Blood Services, Hamilton, Ontario.
W P
WP
Sheffield
P D
PD
Schuyler
M A
MA
Blajchman
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D003001:Cloning, Molecular; D016384:Consensus Sequence; D015698:Genomic Library; D015844:Heparin Cofactor II / Q000096:biosynthesis / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
D011994:Recombinant Proteins; D015844:Heparin Cofactor II