Amplification of the active site of BnLIP3 gene of Brassica napus L. during germination

Lipases are useful enzymes that are responsible for the hydrolysis of triacylglycerides and play an important role in plant growth. In this study, we report a rapid molecular method to amplify a partial sequence of the lipase class 3 family designated BnLIP3 gene of Brassica napus L. in order to follow its expression and analyze its role during seed germination. Therefore, we conceived PCR homologous primers to amplify the active site encoding region of the BnLIP3 family genes. Subsequently, to sequence determination of the 582 bp fragment, we deduced BnLIP3 specific primers for a nested RTPCR application. The deduced 194 amino acid sequence (Genbank 1160264) was found to share 85% of identity with lipase from Arabidopsis thaliana class 3 family. The GxSxG consensus motif near the catalytic triad at the active serine site was also identified. The peptidic sequence showed little homology with mammalian and microbial lipases. RT-PCR analysis indicated that BnLIP3 gene was expressed during B. napus seed germination.Keywords: Brassica napus L., GxSxG lipase, germination, BnLIP3, RT-PCR.African Journal of Biotechnology Vol. 12(25), pp. 3905-391

Identification of Disalicyloyl Curcumin as a Potential DNA Polymerase Inhibitor for Marek’s Disease Herpesvirus: A Computational Study Using Virtual Screening and Molecular Dynamics Simulations

Marek’s disease virus (MDV) is a highly contagious and persistent virus that causes T-lymphoma in chickens, posing a significant threat to the poultry industry despite the availability of vaccines. The emergence of new virulent strains has further intensified the challenge of designing effective antiviral drugs for MDV. In this study, our main objective was to identify novel antiviral phytochemicals through in silico analysis. We employed Alphafold to construct a three-dimensional (3D) structure of the MDV DNA polymerase, a crucial enzyme involved in viral replication. To ensure the accuracy of the structural model, we validated it using tools available at the SAVES server. Subsequently, a diverse dataset containing thousands of compounds, primarily derived from plant sources, was subjected to molecular docking with the MDV DNA polymerase model, utilizing AutoDock software V 4.2. Through comprehensive analysis of the docking results, we identified Disalicyloyl curcumin as a promising drug candidate that exhibited remarkable binding affinity, with a minimum energy of −12.66 Kcal/mol, specifically targeting the DNA polymerase enzyme. To further assess its potential, we performed molecular dynamics simulations, which confirmed the stability of Disalicyloyl curcumin within the MDV system. Experimental validation of its inhibitory activity in vitro can provide substantial support for its effectiveness. The outcomes of our study hold significant implications for the poultry industry, as the discovery of efficient antiviral phytochemicals against MDV could substantially mitigate the economic losses associated with this devastating disease.Funder: King Saud University, Riyadh, Saudi Arabia; Grant(s): RSPD2023R74

Amplification of the active site of BnLIP3 gene of Brassica napus L. during germination

Lipases are useful enzymes that are responsible for the hydrolysis of triacylglycerides and play an important role in plant growth. In this study, we report a rapid molecular method to amplify a partial sequence of the lipase class 3 family designated BnLIP3 gene of Brassica napus L. in order to follow its expression and analyze its role during seed germination. Therefore, we conceived PCR homologous primers to amplify the active site encoding region of the BnLIP3 family genes. Subsequently, to sequence determination of the 582 bp fragment, we deduced BnLIP3 specific primers for a nested RTPCR application. The deduced 194 amino acid sequence (Genbank 1160264) was found to share 85% of identity with lipase from Arabidopsis thaliana class 3 family. The GxSxG consensus motif near the catalytic triad at the active serine site was also identified. The peptidic sequence showed little homology with mammalian and microbial lipases. RT-PCR analysis indicated that BnLIP3 gene was expressed during B. napus seed germination.Glaied Ghram, I.; Belguith, H.; Messaoudi, A.; Fattouch, S.; Vicente Meana, Ó.; Ben Hamida, J. (2013). Amplification of the active site of BnLIP3 gene of Brassica napus L. during germination. African Journal of Biotechnology. 12(25):3905-3913. doi:10.5897/AJB12.2861S39053913122

Three-Dimensional Structure of Lipase Predicted by Homology Modeling Method

Triacylglycerol lipases have been thoroughly characterized in mammals and microorganisms. By contrast, very little is known about plant lipases. In this investigation, a homology model of Arabidopsis thaliana lipase (NP_179126) was constructed using a human gastric lipase (PDB ID: 1HLG), as a template for model building. This model was then assessed for stereochemical quality and side chain environment. Natural substrates: tributyrin, trioctanoin and triolen were docked into the model to investigate ligand-substrate interaction

Classification of EC 3.1.1.3 bacterial true lipases using phylogenetic analysis

Lipases play an important role in lipid metabolism and are produced by a variety of species. All lipases are members of the á/â hydrolase fold super-family. Also, lipases share a conserved active site signature, the Gly-Xaa-Ser-Xaa-Gly motif. To obtain an overview of this industrially and very important class of enzymes and their characteristics, we collected and classified bacterial lipases sequences available from protein databases. Here we proposed an updated and revised classification of family I bacterial "true" lipases based mainly on a comparison of their amino acid sequences and some fundamental physicochemical and biological properties. The result of this work has identified 11 subfamilies of “true” lipases. This work will therefore contribute to a faster identification and to an easier characterization and classification of novel bacterial lipolytic enzymes.Key words: Lipases, phylogenetic analysis, lipolytic enzymes

Identification of Disalicyloyl Curcumin as a Potential DNA Polymerase Inhibitor for Marek's Disease Herpesvirus: A Computational Study Using Virtual Screening and Molecular Dynamics Simulations.

From Europe PMC via Jisc Publications RouterHistory: ppub 2023-09-01, epub 2023-09-12Publication status: PublishedMarek's disease virus (MDV) is a highly contagious and persistent virus that causes T-lymphoma in chickens, posing a significant threat to the poultry industry despite the availability of vaccines. The emergence of new virulent strains has further intensified the challenge of designing effective antiviral drugs for MDV. In this study, our main objective was to identify novel antiviral phytochemicals through in silico analysis. We employed Alphafold to construct a three-dimensional (3D) structure of the MDV DNA polymerase, a crucial enzyme involved in viral replication. To ensure the accuracy of the structural model, we validated it using tools available at the SAVES server. Subsequently, a diverse dataset containing thousands of compounds, primarily derived from plant sources, was subjected to molecular docking with the MDV DNA polymerase model, utilizing AutoDock software V 4.2. Through comprehensive analysis of the docking results, we identified Disalicyloyl curcumin as a promising drug candidate that exhibited remarkable binding affinity, with a minimum energy of -12.66 Kcal/mol, specifically targeting the DNA polymerase enzyme. To further assess its potential, we performed molecular dynamics simulations, which confirmed the stability of Disalicyloyl curcumin within the MDV system. Experimental validation of its inhibitory activity in vitro can provide substantial support for its effectiveness. The outcomes of our study hold significant implications for the poultry industry, as the discovery of efficient antiviral phytochemicals against MDV could substantially mitigate the economic losses associated with this devastating disease