Functional expression, purification, biochemical and biophysical characterizations, and molecular dynamics simulation of a histidine acid phosphatase from Saccharomyces cerevisiae.
World J Microbiol Biotechnol
; 40(6): 171, 2024 Apr 17.
Article
in En
| MEDLINE
| ID: mdl-38630327
ABSTRACT
A histidine acid phosphatase (HAP) (PhySc) with 99.50% protein sequence similarity with PHO5 from Saccharomyces cerevisiae was expressed functionally with the molecular mass of â¼110 kDa through co-expression along with the set of molecular chaperones dnaK, dnaJ, GroESL. The purified HAP illustrated the optimum activity of 28.75 ± 0.39 U/mg at pH 5.5 and 40 ËC. The Km and Kcat values towards calcium phytate were 0.608 ± 0.09 mM and 650.89 ± 3.6 s- 1. The half-lives (T1/2) at 55 and 60 ËC were 2.75 min and 55 s, respectively. The circular dichroism (CD) demonstrated that PhySc includes 30.5, 28.1, 21.3, and 20.1% of random coils, α-Helix, ß-Turns, and ß-Sheet, respectively. The Tm recorded by CD for PhySc was 56.5 ± 0.34ËC. The molecular docking illustrated that His59 and Asp322 act as catalytic residues in the PhySc. MD simulation showed that PhySc at 40 ËC has higher structural stability over those of the temperatures 60 and 80 ËC that support the thermodynamic in vitro investigations. Secondary structure content results obtained from MD simulation indicated that PhySc consists of 34.03, 33.09, 17.5, 12.31, and 3.05% of coil, helix, turn, sheet, and helix310, respectively, which is almost consistent with the experimental results.
Key words
Full text:
1
Database:
MEDLINE
Main subject:
Radioisotopes
/
Saccharomyces cerevisiae Proteins
/
Molecular Dynamics Simulation
/
Magnesium
Language:
En
Journal:
World J Microbiol Biotechnol
Year:
2024
Type:
Article
Affiliation country:
Malaysia