Abstract
Creatine kinases (CKs) are part of the phosphagen kinase (PK) family, a well-studied group of proteins with a variety of naturally occurring quaternary structures (monomers, dimers, fused dimers, and octamers) and high sequence identity (85-99%). All PKs bind phosphagens to catalyze the reversible transfer of a phosphoryl group to adenosine diphosphate, forming adenosine triphosphate and a guanidino (e.g., creatine for creatine kinases, arginine for arginine kinases). It was previously theorized that dimeric CKs evolved from dimeric AKs, which was widely accepted until the discovery of a CK from the protozoan species Perkinsus marinus (PmCK), evoking evolutionary questions since PmCK is the only naturally occurring monomeric CK known. PmCK possesses some of the critical residues structurally important for dimerization in rabbit muscle CK at the location of the hypothesized interface. This suggests that PmCK is either an example of dimeric trait loss or an intermediate between monomeric AKs and dimeric CKs. Site directed mutagenesis was used to form five variants (F178R, D179G, G83D, double (F178R/G83D), and triple (F178R/D179G/G83D)). Previous studies found that variants in an equilibrium of monomers and dimers displayed a large loss of catalytic efficiency, so this study analyzed the kinetics of the monomeric variants (F178R and D179G) to determine if the tertiary or quaternary structure is responsible for this loss of catalytic efficiency. The monomeric variants displayed 2-fold losses of catalytic efficiency, while variants in structural equilibrium displayed 5-fold to 45-fold decreases as compared to the wild type PmCK. The inability to form a stable dimer and loss of catalytic efficiency present in the variants indicate there is a lack of key mutations elsewhere in the protein. This suggests that PmCK’s monomeric structure is not an example of trait loss but is an intermediate between monomeric AKs and dimeric CKs.
Advisor
Snider, Mark
Department
Biochemistry and Molecular Biology
Recommended Citation
Gascoigne, Grace M., "Structural and Kinetic Analysis of Perkinsus Marinus Creatine Kinase: Insight to Quaternary Structure Evolution of Phosphagen Kinases" (2024). Senior Independent Study Theses. Paper 11211.
https://openworks.wooster.edu/independentstudy/11211
Disciplines
Biochemistry | Evolution | Molecular Biology | Structural Biology
Publication Date
2024
Degree Granted
Bachelor of Arts
Document Type
Senior Independent Study Thesis
© Copyright 2024 Grace M. Gascoigne