Effect of mutation in active site residue Trp209 to Val, Leu, Ile and Pro on the catalytic activity and affinity for some benzenesulfonamides of human carbonic anhydrase II

DERYANUR KILIÇ; ORHAN ERDOĞAN; ÖMER İRFAN KÜFREVİOĞLU

Article Details

Effect of mutation in active site residue Trp209 to Val, Leu, Ile and Pro on the catalytic activity and affinity for some benzenesulfonamides of human carbonic anhydrase II

Authors:

DERYANUR KILIÇ | ORHAN ERDOĞAN | ÖMER İRFAN KÜFREVİOĞLU

Publisher:

TUBITAK

Main Topics:

Life sciences

Journal:

Turkish Journal of Biology 2017 Vol.41 Issue 5, pp.935-842

Document Type:

Article

Publication Date:

9/2017

Document Language:

English

Document ID (Org No.):

2000811352

Frequency:

Bimonthly

eISSN:

1303-6092

pISSN:

1300-0152

Abstract:

 

Human carbonic anhydrase II (hCA II) enzyme was firstly expressed using a pET-SUMO expression vector in Escherichia coli and the recombinant enzyme was purified using nickel (Ni2+) affinity chromatography. The substitutions of Trp 209 with four amino acid (Val, Leu, Ile, and Pro) in the hydrophobic pocket of hCA II were conducted using site-directed mutagenesis. The p-nitrophenyl esterase activity of hCA II variants correlates with the hydrophobicity and size of residue, suggesting that the hydrophobic character of this residue is important for catalysis. The Trp 209 was forecast as an important residue and was exposed to computational mutagenesis. This forecast was confirmed experimentally by producing hCA II mutants and determining the resulting affinities towards some benzenesulfonamides. These mutations in the hydrophobic pocket of the enzyme active site decreased the protein expression of hCA II in E. coli, causing the formation of insoluble protein aggregates in many cases. Our findings demonstrated that the Trp 209 in hCA II plays an important role in the folding process and the valine residues are very compatible for the hydrophobic region in the active cavity of this isoenzyme. These mutant proteins will lead to a better understanding of structural functions and drug-based studies in the future.

Article Details

Effect of mutation in active site residue Trp209 to Val, Leu, Ile and Pro on the catalytic activity and affinity for some benzenesulfonamides of human carbonic anhydrase II

Authors: DERYANUR KILIÇ | ORHAN ERDOĞAN | ÖMER İRFAN KÜFREVİOĞLU

Publisher: TUBITAK

Main Topics: Life sciences

Journal: Turkish Journal of Biology 2017 Vol.41 Issue 5, pp.935-842

Document Type: Article

Publication Date: 9/2017

Document Language: English

Document ID (Org No.): 2000811352

Frequency: Bimonthly

eISSN: 1303-6092

pISSN: 1300-0152

Issues

Authors:

DERYANUR KILIÇ | ORHAN ERDOĞAN | ÖMER İRFAN KÜFREVİOĞLU

Publisher:

TUBITAK

Main Topics:

Life sciences

Journal:

Turkish Journal of Biology 2017 Vol.41 Issue 5, pp.935-842

Document Type:

Article

Publication Date:

9/2017

Document Language:

English

Document ID (Org No.):

2000811352

Frequency:

Bimonthly

eISSN:

1303-6092

pISSN:

1300-0152