Scientific Posts
Breaking the Scale Barrier in Biocatalyst Development: AI-Powered Enzyme Engineering for Minimal Variants and Commercial Efficiency - In this study, we developed a sustainable biocatalytic process for sitagliptin production using an AI-driven 6D-grid protein engineering platform. By leveraging molecular interaction data, solvent effects, and a database of 1.39 million structural fragments, we identified high-performing R-transaminase variants while minimizing experimental screening. The engineered enzyme exhibited excellent solubility, stability, and scalability, achieving up to...
Quantum Molecular Dynamics Study on the Reaction Mechanism of Nitrilase toward an Aliphatic Dinitrile Substrate - This study employed quantum mechanical dynamics (QMD) simulations to elucidate the catalytic mechanism of Bacillus safensis nitrilase (BsNIT) toward pentanedinitrile (PD). The results revealed a sequential conversion of PD to 4-cyanobutanoic acid and subsequently to pentanedioic acid through two water-mediated hydrolysis steps involving the catalytic triad (C164, K130, and E38). Key intermediates and transition states...
Reversal of the Leloir pathway to promote galactose and tagatose synthesis from glucose - In this research article, we demonstrate direct conversion of glucose to D-tagatose, a low-calorie rare sugar, using an engineered Escherichia coli whole-cell system. Our study identifies a galactose-1-phosphate phosphatase (DdGal1Pase) with stringent substrate selectivity driven by distinct hydrogen-bond networks, enabling reversal of the Leloir pathway and glucose-based tagatose biosynthesis. This work provides a foundation for...
Unraveling the Structure–Function Relationship and Mechanism of an Important Spiro-Forming Nitrilase Using Metadynamics and Quantum Molecular Dynamics - This study provides the first comprehensive computational investigation of the nitrilase from Bacillus safensis (BsNIT) and Spirosoma linguale DSM 74 (SINIT) for elucidation on the reaction mechanism for nitrile hydrolysis and nitrile hydration reaction respectively. Mechanistic analysis revealed critical catalytic events, including nucleophilic attack by Cys and water-mediated proton transfer by Glu, with covalent substrate...
Multimodal neural network for enhanced protein stability prediction by integration of contact scores and spatial maps - In this study, we developed an AI-driven protein stability prediction framework that integrates AlphaFold-generated structures, molecular refinement, and deep learning techniques. Multiple machine learning and neural network architectures were evaluated, with a multi-input CNN model leveraging protein contact maps delivering the best performance. The framework enables efficient prediction of mutation-induced stability changes, reducing reliance on...
Computational studies on the catalytic potential of the double active site for enzyme engineering - We’re excited to announce our latest publication in Nature Scientific Reports on proteins with double active sites, which could revolutionize enzyme design strategies. We have developed an AI-based tool which creates enzymes with multiple active sites, enhancing their functionality by incorporating diverse catalytic activities. For example, a transaminase enzyme can now also facilitate hydrolysis reactions....
In Silico Screening of Chlorogenic Acids from Plant Sources against Human Translocase-I to Identify Competitive Inhibitors to Treat Diabetes - Chlorogenic acids (CHLs) are known to competitively bind to translocase-I (T1) of the glucose-6-phosphatase (G6 Pase) system, thereby inhibiting the transport of glucose-6-phosphate (G6P). This competitive binding results in a consequential reduction in blood sugar levels. In this study, steered molecular dynamics (SMD) simulation is employed to investigate the interaction between T1 and G6P, aiming...
Towards universal synthetic heterotrophy using a metabolic coordinator - Engineering the utilization of non-native substrates, or synthetic heterotrophy, in proven industrial microbes such as Saccharomyces cerevisiae represents an opportunity to valorize plentiful and renewable sources of carbon and energy as inputs to bioprocesses. We previously demonstrated that activation of the galactose (GAL) regulon, a regulatory structure used by this yeast to coordinate substrate utilization...
Structure, dynamics, and molecular inhibition of the Staphylococcus aureus m1A22-tRNA methyltransferase TrmK - The enzyme m1A22-tRNA methyltransferase (TrmK) from Staphylococcus aureus catalyzes the transfer of a methyl group to the N1 of adenine 22 in bacterial tRNAs. TrmK is essential for S. aureus survival during infection, but has no homolog in mammals, making it a promising target for antibiotic development. Highlights Molecular dynamics simulations point to specific motions...
In-Depth Sequence–Function Characterization Reveals Multiple Pathways To Enhance Enzymatic Activity - Phenyl Ammonia Lyase (PAL) has garnered significant attention as the active ingredient in Pegvaliase, the only FDA-approved drug for treating classical phenylketonuria (PKU). Highlights: Deep Mutational scanning (DMS) to derive functional hotspots Hybrid QM/MM techniques combined with meta-dynamics on all 4 active sites of the PAL tetramer simultaneously, to enhance the sampling of coordinates relevant to...
Extensive Modelling and Quantum Chemical Study of Sterol C-22 Desaturase mechanism: a commercially important Cytochrome P450 family - In this study, we have conducted in-depth modelling studies, extended molecular dynamics experiments, and quantum chemical calculations to throw light on the structure-function relationship and its mechanism of enzyme action. A novel finding on second hydrogen abstraction in the C-22 desaturase mechanism was delineated. Read More: https://doi.org/10.1016/j.cattod.2021.12.004
Identification of a Reaction Intermediate and Mechanism of Action of Intermediary Enzymes in Plumbagin Biosynthetic Pathway - Scaling up is a problem that a lot of biotechnologists find difficult, in terms of manufacturing techniques and products; that’s where we provide biocatalytic solutions at Kcat! The above schema is an intermediary reaction in the Plumbagin Biosynthetic Pathway. We at Kcat have identified two critical steps and respective enzymes in the synthesis of Plumbagin,...
“In Silico Based Engineering Approach to Improve Transaminases for the Conversion of Bulky Substrates” - For more information, refer to https://doi.org/10.1021/acscatal.8b03900
LSFG: Method for capturing atomic details of proteins using a 3d grid for mutational analysis - LSFG is a novel method for engineering proteins by utilizing an atomistic grid-based computational method that analyzes and compares protein atomic compositions. By arranging a protein’s atoms into a fine 3D grid, this approach captures atomic-level details in highly localized regions, allowing comparison of specific protein regions even in the absence of structural similarity. Unlike...
A method for engineering proteins - Presented herein is a method for protein engineering that constructs a three-dimensional grid space around a protein. Different probes simulating amino acid interactions are placed within the grid. Pair Interaction Energy is calculated using the FMO technique for probe-protein interactions. An algorithmic process calculates the sum of PIEs, extending from each grid point to its...
Engineered cytoskeleton actin polypeptide with nitrilase activity - The functional repurposing of structural proteins offers exciting opportunities for sustainable and efficient biocatalysis. This study presents the engineering of two structural proteins, F-actin, to exhibit nitrilase activity, enabling the conversion of 1-(cyanomethyl)cyclohexane-1-carbonitrile into 2-(1-cyanocyclohexyl)acetic acid, a key intermediate in the production of gabapentin. By introducing catalytic residues adapted from nitrilase enzymes, these engineered proteins...
An artificial intelligence method to predict selectivity of transaminases and enantiomeric excess of transaminase-ketone reactions - The present invention describes identifying if a given Transaminase is (R)-Selective or (S)-Selective. Based on the binding of the substrate in the active site of a given transaminase, it is primarily based on torsion, the angle between specific atoms of the cofactor PLP and the catalytic Lysine, and the ten interactions of the substrate with...
Insilico guided crispr-cas9 driven enzyme engineering framework - The invention describes the methods of directing CRISPR complex formation in bacterial cells and utilizing the CRISPR-Cas system to edit a gene of interest in a plasmid. An AI incorporated CRISPR tool to engineer enzymes for better activity and allow cells to undergo specific and random mutation for a variety of research or translational applications,...
A Receptor Dependent-4D QSAR Approach to Predict the Activity of Mutated Enzymes - Screening and selection tools to obtain focused libraries play a key role in successfully engineering enzymes of desired qualities. The quality of screening depends on efficient assays; however, a focused library generated with a priori information plays a major role in effectively identifying the right enzyme. As a proof of concept, for the first time,...
Mechanism of Phytochemicals : Affects the Polymerization and Depolymerisation of Actins - Mechanical understanding of the correlation between actin assembly and ATP hydrolysis has been an object of intensive studies in biochemistry and structural biology. Azadirachtin(A) (AZA), a potential insecticide from neem, binds to actin and induces depolymerization in Drosophila. AZA binds to the pocket the same as that of Latrunculin A (LAT), but LAT inhibits actin...
Engineering Enzymes for Improved Activity at Varying Concentrations of Substrate and Organic Solvent - One of the major problems in the use of enzymes in the industry is their instability under strict process conditions. As the use of organic solvents in industries for hydrophobic substrate solubility or the suppression of water-dependent reactions has become inevitable, the need to engineer enzymes to accommodate the varying solvent conditions and high substrate...
Insight Into Cholinesterase Inhibitors Mechanism By Car–Parrinello Ab Initio Molecular Dynamics In Conjunction With Metadynamics - The current study demonstrates the significant progress made in computational enzymology and exemplifies the power and advantages of employing CPMD simulations in characterizing enzyme mechanisms. Here Acetylcholinesterase inhibitors such as Rivastigmine, Physostigmine, Neostigmine, and Edrophonium binding mechanisms were explored through the metadynamics approach. (In Preparation)
Identification Of Hotspots And Derive Substitution For These Hotspots To Make Enzyme Variants And Screening These Enzyme Variants With Deep Learning Using A-CNN To Derive Top 5 Enzyme Variants With Desired Kinetic Property - The invention is specific to the engineering of enzymes. The Michaelis Complex of E-S reaction is simulated using QM/MM, and MD methods. Using the QM polarized grid maps, MMPBSA. MMGBSA, low energy ES conformations that lead to the formation of attack conformation are identified. Subsequently, the enzyme is designed around the Michaelis complex where the...
Swiss model becomes 25 now! - Congratulations Swissmodel for completing 25 years of service!An excellent and powerful protein modeling server, the first ever modeling server since 2002. Modeled 1000s of protein, cytoskeletal proteins like tubulin, actin, and enzymes for engineering studies like Penicillin G Acylase. A tool that can be used for teaching and as well for research purposes.
Quantum Tunnelling! - Kcat Enzymatic we are working on the similar reactions of Cytochromes as in the Nobel Laureate Dr. Arnold’s lab, however by a different technology that uses Quantum Molecular Dynamics. We wonder if enzymatic reactions take the aid of quantum tunneling effects; specifically, could it be heavy atom tunneling! A quantum mechanical phenomenon where particles penetrate...
Identification of Cryptic Sites/Interactions using Probe-Based Induction Algorithm (MIST)® - Cryptic pockets are sites on enzymes that only become apparent when ligands (substrate, co-substrate, cofactor) binds. They do not correspond to local minima in the computed conformational free energy landscape of the un-liganded proteins. Hence, they close in all the molecular dynamics simulations performed. Their formation is also known to be an interplay between both...
Kcat’s Technology used to derive variants to achieve kg level Substrate conversion - Schematic representation of Kcat’s Technology used to derive variants to achieve kg level substrate conversion with desired regioselectivity, stereospecificity, and enantiomeric excess. A system of Micrometer size with higher enzyme and substrate concentrations at varying pH, temperature, cofactor, co-substrate, and solvent conditions is simulated for microseconds. Information Capturing Technologies:A 7th Dimensional Grid made of Quantum beads that captures...
“7D-QSAR” INTEGRATED WITH QUANTUM MECHANICS A NOVEL TOOL FOR IMPROVING INDUSTRIAL ENZYMES, OUR NEW PATENT - The patented technology works on Quantum mechanics-based probes/grid points, and real solvation using semi-empirical QM simulations. The dynamic probes atoms in the grid are replaced based on the neighbor-neighbor contacts in the conformational transitions. The Dynamic grid points capture high-resolution details of the enzymatic reaction. The method was implemented on an R-specific transaminase enzyme and...
7D GRID ENZYME ENGINEERING ENGINE® - Kcat Enzymatic brings Enzyme Engineering & Biocatalysis Revolution by using upfront in-silico in-house developed methods and algorithms using 7D Grid Technology for enzyme engineering (Patent No: 201841047021). In this line, a new and advanced way of enzyme engineering methods were developed to use enzymes in industrial processes. Our multiscale methods and technologies gives custom engineered... Deep Mutational Scanning Guided Directed Evolution Reveals Multiple Mechanisms to Improve Enzyme Function - This is a collaboratory work with TUFTS University, USA on the advances in Deep mutational scanning to yield hyperactive PAL variants. The reaction mechanism of the enzyme variants was confirmed through QM/MM studies combined with metadynamics. (Under Review)