Prediction of substrate-binding site and elucidation of catalytic residue of a phytase from Bacillus sp.
نویسندگان
چکیده
The present study primarily deals with the identification of substrate-binding site and elucidation of catalytic residue of the phytase from Bacillus sp. (Genbank Accession No. EF536824) employing molecular modeling and site-directed mutagenesis. Homology-based modeling of the Bacillus phytase revealed β-propeller structure with twelve active-site aminoacid residues, viz., D75, R77, Y78, H138, Q140, D189, D190, E191, Y238, Y239, N346 and R348. Docking of substrate Ins(1,2,3,4,5,6)hexakisphosphate with the phytase model disclosed interaction of Y78 residue with the sixth position phosphate, while D75 and R77 residues revealed hydrogen bonding with the fifth position phosphate of the phytate. Analysis of hydrolysis products of phytate indicated the sequential removal of alternate phosphates, resulting in the formation of final product Ins triphosphate. Mutant phytases Y78A/F, derived from site-directed mutagenesis, exhibited complete loss of enzyme activity despite substrate binding, thereby suggesting the intrinsic role of Y78 residue in the catalytic activity. The Bacillus mutant phytases can be used to generate enzyme crystals complexed with phytate and lower Ins phosphates for indepth analysis of substrate binding and catalytic activity of the enzyme.
منابع مشابه
ELUCIDATION OF pK VALUES FOR ACTIVE SITE OF HORSERADISH PEROXIDASE AND BINDING STUDY OF INTERACTION WITH N-PHENYL BENZHYDROXAMIC ACID USING A SPECIAL DIFFERENCE SPECTROPHOTOMETRIC TECHNIQUE
The binding behavior of a competitive inhibitor, N-phenylbenzhydroxamic acid (BHA) against horseradish peroxidase (HRP) was studied in order to understand and predict the interaction mechanism of hydrogen donors with the enzyme. The dissociation constants of the complexes of HRP-BHA, HRP-donor and HRP-BHA-azide were estimated at specified conditions by difference spectroscopy. The binding s...
متن کاملBatch Kinetics and Modeling of Alkaline Protease Production by Isolated Bacillus sp. (RESEARCH NOTE)
The aim of this study was the use of fish waste hydrolysate (FWH) as a substrate for alkaline protease production using isolated Bacillus sp. in a batch system. Then the fermentation kinetics of enzyme production was studied. The results show that with the addition of FWH to the fermentation medium with a final concentration of 4% (optimal concentration), alkaline protease value reached a maxim...
متن کاملPurification and characterization of an acidic, thermophilic phytase from a newly isolated Geobacillus stearothermophilus strain DM12
Microbial phytases were applied mainly to animal and human foodstuffs in order to improvemineral bioavailability and food processing. In addition, phytases have potentialbiotechnological application in various other fields, such as environmental protection,aquaculture and agriculture. Bacillus sp. DM12, an isolate from a hot spring, produces phytase,which catalyzes the hydrolysis of phytic acid...
متن کاملResponse of surface optimization for the enhanced production of alkaline protease isolated from Bacillus sp. with bean husk as a new substrate
Optimization of the fermentation medium for maximum alkaline protease production was carried out. Fifteen positive isolates were examined for their extent of alkaline protease production. The most potent producer was identified as Bacillus sp. The solid substrate screening showed that the combination of wheat straw and bean husk was the best one. The initial screening by using Plackett–Burman’s...
متن کاملBacillus Amyloliquefaciens Alpha-amylase Inhibition by Organic Solvents: A Study on Methanol, Ethanol and Propanol
Alpha-amylase is widely used as an industrial enzyme, and a therapeutic target for which inhibitors are designed. Organic solvents are used to dissolve various compounds that would be studied as moderators of alpha-amylases, but they could themselves affect enzyme activity and stability. Methanol, ethanol and propanol are simple alcohols that may be commonly used to this end, and their effect h...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Enzyme and microbial technology
دوره 51 1 شماره
صفحات -
تاریخ انتشار 2012