Open AccessOptimization of conformational stability and catalytic efficiency in chondroitinase ABC Ι by protein engineering methods
Author(s) -
Shamsi Masoumeh,
Shirdel S. Akram,
Jafarian Vahab,
Jafari S. Shahrbanoo,
Khalifeh Khosrow,
Golestani Abolfazl
Publication year - 2016
Publication title -
engineering in life sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.547
H-Index - 57
eISSN - 1618-2863
pISSN - 1618-0240
DOI - 10.1002/elsc.201600034
Subject(s) - mutant , chemistry , protein engineering , point mutation , enzyme , serine , mutant protein , kinetics , biophysics , catalytic efficiency , protein structure , biochemistry , stereochemistry , biology , physics , quantum mechanics , gene
Chondroitinase ABC Ι can promote the recovery of spinal cord injuries by depolimerization of glycosaminoglycans. However, low thermal stability is one of the limitations regarding its clinical application. In order to increase the conformational stability of the enzyme, Leu 679 at the starting point of a short helix located at the C‐terminal domain of the protein was replaced by serine (L679S mutant) and aspartic acid (L679D mutant). Theoretical and spectroscopic studies showed that the stability of enzyme increased upon mutation. Based on the activity measurements, the catalytic efficiency of L679S was improved in comparison with the wild‐type protein; while that of L679D (a more stabilized protein) was not changed. According to the structural and kinetic data, we proposed a model in which a higher conformational stability results in a slower rate of the formation of the open conformation. On the other hand, a higher flexibility slows down the rate of the formation and holding of the closed conformation. Therefore, the L679S mutant, which is structurally stable relative to the wild‐type protein and is destabilized compared to the L679D mutant, exhibited the best catalytic efficiency. However, it was also found that the L679D mutant was more suitable for long‐term storage of the enzyme.