Characterization of the structural-functional impact of heterozygous missense mutations in genes of the blood coagulation factor XIII that cause mild Factor XIII deficiency

Abstract

The coagulation Factor XIII is a key player in hemostasis that is responsible for the last step of the coagulation cascade in which it covalently cross-links preformed fibrin clots to make them resistant to premature fibrinolysis. The plasma circulating FXIII zymogen is a heterotetrameric complex comprising two catalytic A (FXIIIA₂) and two protective carrier B-Subunits (FXIIIB₂) which are synthesized and secreted into the plasma as homo-dimers from various cell types like monocytes/ macrophages, megakaryocytes, and platelets for the A subunit and hepatocytes for the B subunit. Deficiency of FXIII results in a bleeding predisposition for the individual carrying it. This deficiency can have congenital or acquired origins. The inherited form of FXIII deficiency can be classified into two types based on severity of symptoms: severe and mild FXIII deficiency. While the homozygous inherited form of this deficiency caused by <em>F13A1</em> (OMIM #613225) (FXIIIA subunit) or <em>FXIIIB</em> (OMIM #613235) (FXIIIB subunit) gene mutations is rare (1 in 2-4 million), the milder heterozygous form is more frequent. Only recently, focus has shifted to the mild/heterozygous form of this deficiency that is associated with mild or even an asymptomatic phenotype (unless the affected individual is exposed to some kind of a trauma for e.g. peri-operative settings, accident etc.). Recent investigations from our group in the past five years have shown that inherited mild heterozygous deficiency does have clinical relevance. Identification of heterozygous FXIII deficient patients and extended causality determining research on the related mutations is crucial since the risk of provoked bleeding events (surgery, tooth extraction, trauma) in heterozygous patients can be minimized through early detection. In the last five years, our group has reported 23 mutations from patients with mild FXIII deficiency. Sixteen of these mutations were identified in the <em>F13A1</em> and seven in the <em>F13B</em> gene. In the present study we have performed a comprehensive investigation on the causality of these reported missense mutations using parallel<em> in silico </em>and<em> in vitro</em> approaches to structurally and functionally characterize their underlying pathophysiology. The <em>in vitro</em> methods have been complemented by <em>in silico</em> strategies in which modeling of protein subunits/domains/mutations and simulation/docking based approaches have been applied to explain the in vitro findings. Our analysis shows that these mutations can act on different aspects of FXIII activation and regulation based on the structure functional impact of the particular mutation.