MOLECULAR MECHANISMS OF TYPE-II FACTOR-XIII DEFICIENCY - NOVEL GLY562-ARG MUTATION AND C-TERMINAL TRUNCATION OF THE A SUBUNIT CAUSE FACTOR-XIII DEFICIENCY AS CHARACTERIZED IN A MAMMALIAN EXPRESSION SYSTEM

To explore the biological and clinical implications of the structure/f unction relationships in factor XIII, mutations in two patients with t ype II deficiency were identified and characterized in a mammalian exp ression system, Nucleotide sequence analysis of the A subunit gene sho wed that case no. 1 had a deletion of 4 bp (AATT) in exon XI and that, in case no. 2, Gly562 (GGG) had been replaced by Arg(AGG). The deleti on in case no, 1 leads to a premature termination at codon 464, Restri ction digestion of amplified DNAs confirmed that both cases were homoz ygous for their respective mutations, Reverse transcription-polymerase chain reaction analysis demonstrated that the level of mRNA was great ly reduced in case no, 1, whereas the level of mutant mRNA expressed i n case no, 2 was normal, Molecular modeling calculated that Arg562 cha nged the conformation of the A subunit, suggesting misfolding and/or d estabilization of the molecule. To determine how these mutations impai red synthesis of the A subunit, recombinant A subunits bearing the mut ations were expressed in mammalian cells. Pulse-chase experiments show ed that the mutants were synthesized normally but disappeared rapidly, whereas the wild-type remained. These results indicate that both muta nt proteins with an altered conformation become prone to rapid degrada tion, resulting in factor XIII deficiency in these patients. (C) 1998 by The American Society of Hematology.