کد مقاله کد نشریه سال انتشار مقاله انگلیسی نسخه تمام متن
4752733 1416366 2017 7 صفحه PDF دانلود رایگان
عنوان انگلیسی مقاله ISI
Research PaperImprovement of activity and stability of Chondroitinase ABC I by introducing an aromatic cluster at the surface of protein
موضوعات مرتبط
مهندسی و علوم پایه مهندسی شیمی بیو مهندسی (مهندسی زیستی)
پیش نمایش صفحه اول مقاله
Research PaperImprovement of activity and stability of Chondroitinase ABC I by introducing an aromatic cluster at the surface of protein
چکیده انگلیسی

Chondroitinase ABC I (ChABC I) has been shown to depolymerize a variety of glycosaminoglycan substrates and promote regeneration of damaged spinal cord. However, to date, intrathecal delivery methods have been suboptimal largely due to enzyme instability which necessitates repeated administration to the injured loci. Among the aromatic amino acids, tyrosine has been shown to be more effective in creation of stable clusters and further stabilize of the proteins. Bioinformatics approaches have been used to examine the effect of an extra aromatic cluster at the surface of ChABC I. In this study two amino acids i.e., Asn806 and Gln810 were mutated to tyrosine and to alanine as negative control. In this way, four variants i.e., N806Y/Q810Y, N806A/Q810Y, N806Y/Q810A and N806A/Q810A were created. The results showed that N806Y/Q810Y mutation improved both activity and thermal stability of the enzyme while Ala substitution reduced the enzyme activity and destabilized it. Structural analysis of mutants showed an increase in intrinsic fluorescence intensity and secondary structure content of N806Y/Q810Y mutant when compared to the wild type enzyme indicating a more rigid structure of this variant. Moreover, the N806Y/Q810Y enzyme displayed a remarkable resistance against trypsin degradation with a half-life (t1/2) of 45.0 min versus 32.5 min of wild-type. In conclusion, the data revealed that structural features and activity of ChABC I can be improved by introducing appropriate aromatic clusters at the surface of the enzyme.

ناشر
Database: Elsevier - ScienceDirect (ساینس دایرکت)
Journal: Enzyme and Microbial Technology - Volume 105, October 2017, Pages 38-44
نویسندگان
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