In-silico identification of lysine residue for lysozyme immobilization on dialdehyde cellulose.

In the realm of enzymes, the Enzyme Immobilization technique can be extremely beneficial. More research into computational approaches could lead to a better understanding as well as lead us in the direction of a more environmentally friendly and greener path. In this study, molecular modelling techniques were used to collect information regarding the immobilization of Lysozyme (EC 3.2.1.17) on Dialdehyde Cellulose (CDA). Lysine, being the most nucleophilic, is most likely to interact with dialdehyde cellulose. Enzyme substrate interactions have been studied with and without the refinement of modified lysozyme molecules. A total of six CDA-modified lysine residues were selected for the study. The docking process for all modified lysozymes was carried out using four distinct docking programs: Autodock Vina, GOLD, Swissdock, and iGemdock. The binding affinity (-7.8 & -8.0 kcal mol-1 in case of non-refinement and -4.7 & -5.0 kcal mol-1 in case of refinement), calculated from Autodock vina, as well as the interaction similarity of Lys116 immobilized lysozyme with its substrate, were found to be 75 % (W/o simulation) & 66.7 % (With simulation) identical with the reference case (unmodified lysozyme) if Lys116 is bound to Dialdehyde Cellulose. The approach described here is utilized to identify amino acid residues that are used in the immobilization of lysozyme.

Cellulose tosylate as support for α-amylase immobilization.

Tremendous potential exists to use cellulose as a support for the immobilization of enzymes. In the current study, cellulose was transformed into cellulose tosylate, and α-Amylase was immobilized using the derivatized polymer. Techniques like Fourier transform infrared, scanning electron microscopy, thermogravimetric analysis, and X-ray diffraction methods were used to characterize the support. The support is a perfect illustration of how both covalent and hydrophobic/ionic types of immobilizations can be experimentally used on support. The support was found to show max degradation at 320.2 °C with 3.2 % residual substance and crystallinity of about 56.6 %. The support presented maximum enzyme loading at the support: enzyme ratio of 1:4. The immobilized enzyme displayed two ideal pH values (about 4.6 and 7) and two ideal temperatures (approximately 60 °C & 40 °C). It was discovered that the immobilized α-amylase could be used easily eight times with 9.9 % residual activity. The findings of this study show that the immobilized cellulose tosylate enzyme has the potential for application in both acidic and neutral pH environments with the best activity for commercial use.

In-silico identification of lysine residue for α-Amylase immobilization on dialdehyde cellulose.

Enzymes are the precious gift of nature to humans. The wise utilization of enzymes may reduce energy needs of humans and the Immobilization technique can help a lot in this regard. This aspect overcomes limitations of the enzymes, therefore providing an opportunity to explore enzymatic chemistry further. In the present context, it is quite cumbersome & costly to identify the amino acid of enzymes involved in the covalent mode of Immobilization. In the present study, molecular modeling techniques were used to do this difficult task. The present work used molecular modeling methods to extract information about the immobilization of α-Amylase (E.C.3.2.1.1) on Dialdehyde Cellulose. The Lysine residue is the most probable residue to interact with Dialdehyde Cellulose. In the present work, a total of 23 lysine residues were used to study covalent binding behavior with α-Amylase. It was found that if Lys142 is involved in binding with Dialdehyde Cellulose then binding affinity (-6.1 & -5.9 kcal mol-1), as well as the involvement of amino acids of both free α-Amylase and Lys142 immobilized α-Amylase with the starch substrate, were found to be similar. The technique reported here is used for the identification of amino acid residue for the Immobilization of enzymes.

Comparative study of covalent and hydrophobic interactions for α-amylase immobilization on cellulose derivatives.

Indispensability of enzymes in living systems, their unique characteristics and simultaneous focus on development of greener methods have led to substitution of various chemical reactions by enzyme catalyzed reactions. One of the aspects in enzyme research is immobilization of enzymes. Immobilization provides a platform for reusability of significant enzymes. Varieties of methods have been explored for enzyme immobilization such as entrapment, adsorption, ionic interactions etc. Keeping in view the industrial utility of α-Amylase in leather, paper and other industries related to starch hydrolysis, we immobilized α-Amylase on cellulose isolated from banana peel. In present study, two different methods of immobilization - covalent bonding (Cellulose Dialdehyde as a support) and hydrophobic interactions (Nano Cellulose- Cetyl Trimethyl Ammonium Bromide) were used. Cellulose obtained from bio-waste has been characterized using Fourier transform Infrared Spectroscopy (FT-IR), Thermogravimetric Analysis (TGA), Scanning Electron Microscopy (SEM), X-ray Diffraction (XRD). In this comparative study, Cellulose Dialdehyde (CDA) immobilized enzyme depicts high reusability, good enzyme loading, storage capacity up to 49 days, optimum pH 6, optimum temperature 95 °C, good pH and thermal stability as compared to native enzyme having optimum pH and temperature of 7 and 37 °C. On the contrary, nanocellulose - Cetyl Trimethyl Ammonium Bromide (NC-CTAB) matrix shows good enzyme loading and optimum pH shift of about 3 units but poor recyclability. Outcome of this study presents the promising nature of covalent mode of immobilization for industrial use.

Insight from the structural molecular model of cytidylate kinase from Mycobacterium tuberculosis.

Mycobacterium tuberculosis is a gram-positive bacterium causes tuberculosis in human. H37Rv strain is a pathogenic strain utilized for tuberculosis research. The cytidylate mono-phosphate (CMP) kinase of Mycobacterium tuberculosis belongs to the family nucleoside mono-phosphate kinase (NMK), this enzyme is required for the bacterial growth. Therefore, it is important to study the structural and functional features of this enzyme in the control of the disease. Hence, we developed the structural molecular model of the CMP kinase protein from Mycobacterium tuberculosis by homology modeling using the software MODELLER (9v10). Based on sequence similarity with protein of known structure (template) of Mycobacterium smegmatis (PDB ID: 3R20) was chosen from protein databank (PDB) by using BLASTp. The energy of constructed models was minimized and the qualities of the models were evaluated by PROCHECK and VERRIFY-3D. Resulted Ramachandran plot analysis showed that conformations for 100.00% of amino acids residues are within the most favored regions. A possible homologous deep cleft active site was identified in the Model using CASTp program. Amino acid composition and polarity of that protein was observed by CLC-Protein Workbench tool. Expasy's Prot-param server and CYC_REC tool were used for physiochemical and functional characterization of the protein. Studied of secondary structure of that protein was carried out by computational program, ProFunc. The structure is finally submitted in Protein Model Database. The predicted model permits initial inferences about the unexplored 3D structure of the CMP kinase and may be promote in relational designing of molecules for structure-function studies.