Factor XIII is found in quite different locations including plasma (A2B2), the cellular (A2) of platelets, megakaryocytes, and monocytes, and (A2) in bone extracellular matrix. Plasma FXIII A2B2 is activated proteolytically by thrombin and low mM Ca2+ whereas FXIII A2 is activated nonproteolytically by a range of Ca2+ concentrations. The resultant transglutaminases (FXIII A* and A) introduce covalent ?-glutamyl-?-lysyl crosslinks into protein networks. The main objective of our research is to decipher the kinetic and conformational differences between the two activated forms of Factor XIII (FXIII) and to characterize their ability to affect fibrin clot architecture and related processes. FXIII has roles in hemostasis, venous thrombosis, cardiac repair, bone remodeling, innate immunity, etc. There are, however, few examples of strategies to therapeutically target FXIII. To further enhance such developments, critical gaps in knowledge need to be addressed. We need to know more about 1) the substrate specificities and structural features of activated FXIII species from different activation pathways and 2) how fibrin clot architecture is controlled by FXIII catalyzed crosslinking reactions involving the fibrin ?C region and ? module. The proposed research will thus address two hypotheses: 1) Factor XIII is hypothesized to have optimized activation pathways, protein structural features, and substrate specificities to effectively target its actions in different physiological environments. Research will focus a) on the kinetic properties of proteolytic FXIII A* and nonproteolytic FXIII A, b) on probing conformational roles of FXIII Activation Peptide interactions, and c) on enhancing calcium binding. 2) The fibrin(ogen) ?C region and ? module are hypothesized to enhance their interactions with FXIII by utilizing distinct reactive glutamine environments and employing binding segments for FXIII. a) With ?C (233-425), we will focus on quantitatively exploring how individual reactive Qs will facilitate the reactivities of remaining Qs. b) With ?-module (142-411), we will rank the reactivities of Q398/Q399 and assess the role that anchoring sites plays in controlling substrate specificity. The biochemical methods required to address these aims include activity assays, fibrin crosslinking gels, mass spectrometry, solution NMR, and analytical ultracentrifugation. Our contributions to this research field will have a positive impact because they will help to address unresolved issues associated with FXIII structure-function, and as a result, lead to new innovative strategies to manage FXIII in particular physiological locations

Public Health Relevance

Factor XIII has been associated with cardiovascular disease, bleeding disorders, wound healing, innate immunity, and bone dynamics. Using innovative approaches, the proposed research will provide critical information on how to control the ability of Factor XIII to introduce rigid crosslinks into fibrin blood clots and other physiological environments. The research proposed is relevant to public health because its discoveries may lead to the development of novel therapeutic agents to address such medical conditions. Successes in the research projects will support the NIH mission to increase biomedical knowledge and to reduce human disease and disability.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Academic Research Enhancement Awards (AREA) (R15)
Project #
2R15HL120068-03
Application #
9811889
Study Section
Special Emphasis Panel (ZRG1)
Program Officer
Warren, Ronald Q
Project Start
2013-08-05
Project End
2022-07-31
Budget Start
2019-08-01
Budget End
2022-07-31
Support Year
3
Fiscal Year
2019
Total Cost
Indirect Cost
Name
University of Louisville
Department
Chemistry
Type
Schools of Arts and Sciences
DUNS #
057588857
City
Louisville
State
KY
Country
United States
Zip Code
40292
Anokhin, Boris A; Stribinskis, Vilius; Dean, William L et al. (2017) Activation of factor XIII is accompanied by a change in oligomerization state. FEBS J 284:3849-3861
Billur, Ramya; Ban, David; Sabo, T Michael et al. (2017) Deciphering Conformational Changes Associated with the Maturation of Thrombin Anion Binding Exosite I. Biochemistry 56:6343-6354
Jadhav, Madhavi A; Goldsberry, Whitney N; Zink, Sara E et al. (2017) Screening cleavage of Factor XIII V34X Activation Peptides by thrombin mutants: A strategy for controlling fibrin architecture. Biochim Biophys Acta Proteins Proteom 1865:1246-1254
Mouapi, Kelly Njine; Bell, Jacob D; Smith, Kerrie A et al. (2016) Ranking reactive glutamines in the fibrinogen ?C region that are targeted by blood coagulant factor XIII. Blood 127:2241-8
Doiphode, Prakash G; Malovichko, Marina V; Mouapi, Kelly Njine et al. (2014) Evaluating factor XIII specificity for glutamine-containing substrates using a matrix-assisted laser desorption/ionization time-of-flight mass spectrometry assay. Anal Biochem 457:74-84