MOLECULAR AND CELLULAR BASIS FOR TYPE-I HEPARIN-COFACTOR-II DEFICIENCY (HEPARIN-COFACTOR-I AWAJI)

Heparin cofactor II (HCII) is a serine proteinase inhibitor in human p lasma that rapidly inhibits thrombin in the presence of dermatan sulfa te or heparin. To understand the molecular mechanism for HCII deficien cy in a patient with reduced circulating HCII antigen, we studied a Ja panese patient with type I HCII deficiency who suffered from angina pe ctoris and coronary artery disease, Polymerase chain reaction (PCR)-ba sed sequence analysis showed that the propositus' gene for HCII (HCII Awaji gene) had a thymine insertion after codon (GAT) for Asp88 in exo n II, resulting in a frameshift mutation. Consequently, the abnormal H CII Awaji protein was suggested to have an altered amino acid sequence from position 89 and terminate at 107, thus being composed of the NH2 -terminal one fifth of normal HCII and dysfunctional for thrombin inhi bition, The molecular weight and pI value of HCII Awaji were calculate d to be 12,040 and 3.6, respectively, without posttranslational modifi cation. Mutagenic PCR followed by the Tsp5091 digestion showed that a half of the PCR products derived from the propositus and his sister wa s cleaved, suggesting that his sister also has the same mutant allele, Crossed-immunoelectrophoresis and Western blot analyses of plasma and urine from the propositus and of plasma from his sister did not provi de evidence for the existence of the abnormal HCII, suggesting that li ttle truncated HCII was circulating in the patient's blood. However, s table expression assay using human kidney 293 cells transfected with t he expression vector containing cDNA encoding wild-type or Awaji-type HCII showed that mutant as well as wildtype HCII was secreted into cul ture medium normally, These results suggest that the abnormal HCII Awa ji protein is secreted normally, but rapidly degraded in the circulati ng blood. (C) 1996 by The American Society of Hematology.