Identification and computationally-based structural interpretation of naturally occurring variants of human protein C

Abstract

Protein C (PC) is a key regulator of blood clotting and inflammation. Its inherited deficiency is associated with venous thromboembolism, and recombinant activated PC is currently used to increase survival in severe sepsis. The molecular basis of inherited PC deficiency is heterogeneous. Due to its multiple physiologic interactions and functions, and its modular structure, natural variants aid in the understanding of the relationship between critical residues and discrete functions. This knowledge has important therapeutic implications in the planning of a recombinant activated PC with a specific therapeutic target and devoid of major collateral effects. A way of predicting important functional consequences of residue variation is the use of molecular modeling and structural interpretation of amino acidic substitutions. A study of 21 out of 32 identified PC gene (PROC) variants is presented. For three of them, localized in the active site, electrostatic potential variation was calculated. For more than half of the studied variants, an explanation for the functional impairment could be derived from computational analysis, allowing a focused choice of which variants it is worthwhile pursuing.

Genetic characterization of antithrombin, protein C and protein S deficiencies in Polish patients

“…These dimers may accumulate intracellularly in hepatocytes, affecting the synthesis of a wild -type AT and other inflammatory or mutations at the same position, p.Gln226His and p.Gln226Leu, have been described in a newborn Spanish child with purpura fulminans and a Spanish patient screened for thrombophilia after a DVT event, respectively. 37, 38 In the current study, the missense variant Gln226Leu in exon 7 was also detected in a 49 -year -old male patient with DVT. Interestingly, the p.Cys106Arg variant, previously described by us in a young Polish man with provoked DVT and PE, 39 was detected in the current study in 3 other unrelated patients.…”

Compound heterozygous protein C deficiency in a family with venous thrombosis: Identification and in vitro study of p.Asp297His and p.Val420Leu mutations

“…Interestingly, several mutations near the Val420 residue have been reported to cause type II PCD, i.e. p.Trp422Cys, p.Trp422Gly, p.Gly423Ser, and p.Gly423Asp (Marchetti et al, 1993;Miyata et al, 1996;Levo et al, 2000;Rovida et al, 2007). Moreover, the heterozygotes of p.Gly423Ser also showed PC:A reduced to 58-65% (amidolytic activity) and 52-59% (clotting activity), and higher PC:Ag (144-156%).…”

“…According to molecular modeling, amino acid Trp422 is located at the P1 specificity substrate-binding pocket of the serine protease and the Ser-Trp-Gly triplet therein is conserved in most serine proteases (Miyata et al, 1996). Mutations altering the conformation of the pocket affected the substrate recognition site and prevented the correct presentation of the scissile peptide bond to the catalytic residue (Rovida et al, 2007). We assumed that p.Val420Leu may lead to the type II deficiency by the same mechanism.…”

ProCMD: a database and 3D web resource for protein C mutants

“…Of these, a set of 21 variants were identified by our group through the screening of the protein C gene of 42 patients with a phenotypic functional deficiency of protein C [17]. The remaining portion of the dataset consists of missense and nonsense variants already reported, obtained from 3 different sources: the database of mutations of protein C gene [10,11], the Human Gene Mutation Database [12,13] and the Swiss-Prot Variant Page [14,15].…”

“…It includes the description of 21 new variants that we have identified and analyzed in a previous work [16][17]. The database is integrated with an interactive search interface and with tools for structure visualization and mutant modelling.…”