Investigating Activity Loss in an Engineered Monomeric Creatine Kinase through the Lens of a Sponge-derived CK – An Exploration of Quaternary Structure Significance

Authors

Willow H. Reddish, The College of WoosterFollow

Abstract

Investigating quaternary structure in proteins, particularly in the context of phosphagen kinases, provides valuable insights into protein evolution and its applications. This study focuses on creatine kinase, a member of this enzyme family which displays diverse quaternary structures but appears in nature almost exclusively as a dimer. The question arises: why do creatine kinases, and proteins in general, tend to form dimers? Previous research on rabbit muscle creatine kinase suggested that dimerization is crucial for maintaining activity, potentially due to obligate cooperativity. To test this theory, this study mutated two residues in the non-cooperative creatine kinase from Tethya aurantia (R148 and D210) to alanine, which are fully conserved in phosphagen kinase dimers and therefore believed to hold significance in the formation of quaternary structure. The R148A variant successfully formed a monomer. The monomeric R148A CK showed a 2-fold reduction in catalytic activity and a 3-fold reduction in efficiency, suggesting an insignificant loss of activity. The D210A variant remained a dimer and remarkably lost more activity and efficiency than that of the R148A monomer. Key conclusions of the current study include the critical role of Arg148 in dimerization and the potential impact of Asp210 on catalysis and conformational changes that may be critical for active site organization. The broader implication is that dimerization may not be essential for enzyme activity in non-cooperative phosphagen kinases, suggesting that dimerization in creatine kinases lacking cooperativity might have evolved for stability, but the reason behind dimerization in the TaCK remains unclear.

Advisor

Snider, Mark

Department

Biochemistry and Molecular Biology

Recommended Citation

Reddish, Willow H., "Investigating Activity Loss in an Engineered Monomeric Creatine Kinase through the Lens of a Sponge-derived CK – An Exploration of Quaternary Structure Significance" (2024). Senior Independent Study Theses. Paper 11193.
https://openworks.wooster.edu/independentstudy/11193

Keywords

phosphagen kinase, creatine kinase, quaternary structure, dimer, dimerization, site directed mutagenesis, Tethya aurantia

Publication Date

2024

Degree Granted

Bachelor of Arts

Document Type

Senior Independent Study Thesis