title: Design of Factor XIII V34X activation peptides to control ability to interact with thrombin mutants.
abstract: Thrombin helps to activate Factor XIII (FXIII) by hydrolyzing the R37-G38 peptide bond. The resultant transglutaminase introduces cross-links into the fibrin clot. With the development of therapeutic coagulation factors, there is a need to better understand interactions involving FXIII. Such knowledge will help predict ability to activate FXIII and thus ability to promote/hinder the generation of transglutaminase activity. Kinetic parameters have been determined for a series of thrombin species hydrolyzing the FXIII (28-41) V34X activation peptides (V34, V34L, V34F, and V34P). The V34P substitution introduces PAR4 character into the FXIII, and the V34F exhibits important similarities to the cardioprotective V34L. FXIII activation peptides containing V34, V34L, or V34P could each be accommodated by alanine mutants of thrombin lacking either the W60d or Y60a residue in the 60-insertion loop. By contrast, FXIII V34F AP could be cleaved by thrombin W60dA but not by Y60aA. FXIII V34P is highly reliant on the thrombin W215 platform for its strong substrate properties whereas FXIII V34F AP becomes the first segment that can maintain its K(m) upon loss of the critical thrombin W215 residue. Interestingly, FXIII V34F AP could also be readily accommodated by thrombin L99A and E217A. Hydrolysis of FXIII V34F AP by thrombin W217A/E217A (WE) was similar to that of FXIII V34L AP whereas WE could not effectively cleave FXIII V34P AP. FXIII V34F and V34P AP show promise for designing FXIII activation systems that are either tolerant of or greatly hindered by the presence of anticoagulant thrombins.
authors.affiliation: Chemistry Department, University of Louisville, Louisville, KY, USA.
authors.forename: Madhavi A
authors.initials: MA
authors.lastname: Jadhav
authors.forename: R Cory
authors.initials: RC
authors.lastname: Lucas
authors.forename: Whitney N
authors.initials: WN
authors.lastname: Goldsberry
authors.forename: Muriel C
authors.initials: MC
authors.lastname: Maurer
mesh_terms: D019943:Amino Acid Substitution; D000925:Anticoagulants / Q000737:chemistry / Q000378:metabolism; D015195:Drug Design; D004353:Drug Evaluation, Preclinical; D005176:Factor XIII / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry / Q000378:metabolism; D010446:Peptide Fragments / Q000138:chemical synthesis* / Q000378:metabolism / Q000494:pharmacology*; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D013917:Thrombin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014633:Valine / Q000235:genetics
chemical_list: D000925:Anticoagulants; D050505:Mutant Proteins; D010446:Peptide Fragments; D005176:Factor XIII; D013917:Thrombin; D014633:Valine