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Effect of Allosteric Changes in MERS 3CL protease Enzymatic Activity and Dimerization
Author(s) -
Gonzalez Leticia Sofia,
Anson Brandon,
Mesecar Andrew
Publication year - 2017
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.31.1_supplement.601.7
Subject(s) - allosteric regulation , mutagenesis , protease , cysteine , enzyme , binding site , biochemistry , biology , conserved sequence , mutant , chemistry , peptide sequence , gene
MERS‐CoV, the Middle Eastern Respiratory Syndrome Coranavirus, was first identified in late 2012 causing public health havoc as more research tried to identity its virulence and relationship to SARS‐CoV. Currently, MERS‐CoV appears to have a 35% fatality rate for those infected. The catalytic dyad in its 3CLprotease (cysteine and histidine) is conserved within other beta coronaviruses. Overall there is ~98% conservation of the MERS‐CoV, HKU4, HKU5 proteome, yet the three viruses vary significantly in their dimerization binding affinity of their 3Clproteases. HKU4, HKU5 appear to have much stronger binding of their 3CLproteases, while MERS's 3CLprotease binding affinity constant is significantly lower. With a large percent of conserved proteomics it is crucial to further understand why the small percent of non‐conserved protein regions within the 3CLproteases contribute to great differences in affinity constant for MERS 3CLpro. It was the goal of the project is to identify allosteric interactions that would make a stronger dimer within the MERS‐CoV 3CLprotease using and mutating non‐conserved regions in the 3CLproteases family. Through site directed mutagenesis a A15N mutant was cloned, purified and its kinetic and binding affinity was tested. FRET‐based kinetic assays for A15N show that A15N 3CLpro binding affinity was weaker as compared to 3CL protease wild‐type. This results leaves further opportunities to explore other important sites (amino acids/bonds) within the dimer interface that influence more heavily to its dimerization. Support or Funding Information National Science Foundation