Date on Senior Honors Thesis

5-2019

Document Type

Senior Honors Thesis

Degree Name

B.S.

Department

Chemistry

Degree Program

College of Arts and Sciences

Author's Keywords

FXIII; alphaC; fibrinogen; glutamines; clotting; blood

Abstract

The reaction between transglutaminase factor XIII (FXIII) and fibrin is a key process in the final stages of blood coagulation. The αC region of fibrin contains three reactive glutamines (Q237, Q328, and Q366) which are crosslinked by FXIII, as well as a FXIII anchoring site (E396). Previous work has helped elucidate the FXIII-αC interaction, but much is still unknown about the key residues in the αC region.

Four crosslinking assays were carried out to characterize the contribution of the αC anchoring site and reactive glutamines in crosslinking by FXIII. The effects of individual E396A, Q366N, and Q328P (Fibrinogen Seoul II mutant) mutations in Fbg αC were monitored through a GEE crosslinking/ MALDI-TOF MS strategy. These studies revealed that, at physiological conditions, the E396 anchoring site plays the largest role in crosslinking of the most reactive glutamine, Q237, while the other two glutamines had minimal impacts. In addition, the two less reactive glutamines, Q328 and Q366, appear to compete with each other as substrates for FXIII.

Lay Summary

The final steps of blood coagulation involve the formation of a hard clot. This clot is formed through crosslinks between glutamine (Q) and lysine (K) amino acids in the fibrin units by the enzyme Factor XIII (FXIII). One fibrin region where these crosslinks are introduced is the αC region, containing three reactive glutamines, (Q237, Q366, and Q328) and an anchoring site for FXIII containing the amino acid glutamate (E396). Q237 is the most reactive glutamine, and was the primary focus of the project. The overall goal was to investigate how certain mutations to these amino acids would influence fibrin clot properties. Three mutations (E396A, Q366N, and Q328P) were used to assess how losses of key amino acids would affect FXIII dependent crosslinking. Results suggested that, at physiological conditions, the E396 anchoring site plays the largest role in aiding crosslinking in the αC region, but not as much as was previously thought. In addition, the two less reactive glutamines, Q328 and Q366, may be in competition with each other to be crosslinked.

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