MOLECULAR CLONING AND PROTEIN MODELING CONFIRM PRESENCE OF AMINOCYCLOPROPANE-1-CARBOXYLATE (ACC) DEAMINASE GENE IN Bacillus subtilis EG21 STRAIN
Abstract
The aim of this work was to confirm the presence of ACC deaminase gene in the plant growth promoting and salt-stress ameliorating bacterial EG21 strain isolated from cotton plants in salt affected soil. TheEG21 Strain was qualitatively evaluated for its ACC deaminase activity. The ACC deaminase gene (acdS) was amplified from extracted genomic makeup and cloned into pCR™4-TOPO® plasmid cloning vector. The recombinant plasmids were used to transform E. coli competent cells which were screened for successful transformants. Recombinant plasmids were recovered and used for sequence analysis. The obtained sequence of the acdS gene as well as the deduced amino acid sequence of the corresponding protein were analyzed using the Basic Local Alignment Search Tool (BLAST) that available in the GenBank databases for nucleotide and protein sequences. Bioinformatic tools were used to model the acdS gene. Finally, the 3D structure of the modeled protein had met and satisfied the requested threshold for model quality and reliability.
References
Alfiky, A., L’Haridon F., Abou-Mansour E. and Weisskopf L., (2022). isease Inhibiting Effect of Strain Bacillus subtilis EG21 and Its Metabolites Against Potato Pathogens Phytophthora infestans and Rhizoctonia solani. Phytopathology®. https://doi.org/10.1094/phyto-12-21-0530-r
Anand G., Bhattacharjee A., Shrivas V. L., Dubey S. and Sharma S., (2021). ACC deaminase positive Enterobacter-mediated mitigation of salinity stress, and plant growth promotion of Cajanus cajan: a lab to field study. Physiol. Mol. Biol. Plants 27, 1547-1557. https://doi.org/10.1007/s12298-021-01031-0
Benkert P., Biasini M. and Schwede T., (2011). Toward the estimation of the absolute quality of individual protein structure models. Bioinformatics, 27: 343-350. https://doi.org/10.1093/bioinformatics/btq662
Biasini M., Bienert S., Waterhouse A., Arnold K., Stude, G., Schmidt T., Kiefer F., Cassarino T. G., Bertoni M., Bordoli L. and Schwede T., (2014). SWISS-MODEL: Modelling protein tertiary and quaternary structure using evolutionary information. Nucleic Acids Res. 42: 252-258. https://doi.org/10.1093/nar/gku340
Chanratana M., Joe M. M., Roy Choudhury A., Anandham R., Krishnamoorthy R., Kim K., Jeon S., Choi Joonho Choi and Jeongyun Sa T., (2019). Physiological response of tomato plant to chitosan-immobilized aggregated Methylobacterium oryzae CBMB20 inoculation under salinity stress. 3 Biotech 9, 1-13. https://doi.org/10.1007/s13205-019-1923-1
Checcucci A., Azzarello E., Bazzicalupo M., Carlo A. De, Emiliani G., Mancuso S., Spini G., Viti C. and Mengoni A., (2017). Role and regulation of ACC deaminase gene in Sinorhizobium melilotr: Is it a symbiotic, rhizospheric or endophytic gene? Front. Genet. 8. https://doi.org/10.3389/fgene.2017.00006
Conde-Avila V., Ortega-Martínez L. D., Loera O., Pérez-Armendáriz B. and Martínez Valenzuela C., (2022).
Encapsulation of Azotobacter vinelandii ATCC 12837 in Alginate-Na Beads as a Tomato Seedling Inoculant. Curr. Microbiol. 79: 1-10. https://doi.org/10.1007/s00284-022-02797-6
Fatma M., Asgher M., Iqbal N., Rasheed F., Sehar Z., Sofo A. and Khan N. A., (2022). Ethylene Signaling under Stressful Environments: Analyzing Collaborative Knowledge. Plants, 11: 1-29. https://doi.org/10.3390/plants11172211
Gowtham H. G., S., B. S., M., M., N., S., Prasad M., Aiyaz M., K. N., A., S. R. N., (2020). Induction of drought tolerance in tomato upon the application of ACC deaminase producing plant growth promoting rhizobacterium Bacillus subtilis Rhizo SF 48. Microbiol. Res. 234, 126422. https://doi.org/10.1016/j.micres.2020.126422
Gupta S. and Pandey S., (2019). Unravelling the biochemistry and genetics of ACC deaminase-An enzyme alleviating the biotic and abiotic stress in plants. Plant Gene 18: 100175. https://doi.org/https://doi.org/10.1016/j.plgene.2019.100175
Horn D., (2005). Directional enrichment of directly cloned PCR products. Biotechniques 39, 40-46. https://doi.org/10.2144/05391BM03
Jha B., Gontia I. and Hartmann A., (2012). The roots of the halophyte Salicornia brachiata are a source of new halotolerant diazotrophic bacteria with plant growth-promoting potential. Plant Soil 356: 265-277. https://doi.org/10.1007/s11104-011-0877-9
Jung B.K., Ibal J. C., Pham H. Q., Kim M. C., Park G. S., Hong S. J., Jo H. W., Park C. E., Choi S. D., Jung Y., Tagele S. B. and Shin J. H., (2020). Quorum Sensing System Affects the Plant Growth Promotion Traits of Serratia fonticola GS2. Front. Microbiol., 11: 1-12. https://doi.org/10.3389/fmicb.2020.536865
Lamaoui M., Jemo, M., Datla R. and Bekkaoui F., (2018). Heat and drought stresses in crops and approaches for their mitigation. Front. Chem. 6: 1-14. https://doi.org/10.3389/fchem.2018.00026
Ma W., Charles T. C. and Glick B. R., (2004). Expression of an exogenous 1-aminocyclopropane-1-carboxylate deaminase gene in Sinorhizobium meliloti increases its ability to nodulate alfalfa. Appl. Environ. Microbiol., 70: 5891- 5897. https://doi.org/10.1128/AEM.70.10.5891-5897.2004
Ma W., Sebestianova S. B., Sebestian J., Burd G. I., Guinel F. C. and Glick B. R., (2003). Prevalence of 1-aminocyclopropane-1-carboxylate deaminase in Rhizobium spp. Antonie van Leeuwenhoek, Int. J. Gen. Mol. Microbiol. 83, 285-291. https://doi.org/10.1023/A:1023360919140
Naing A. H., Maung T. T. and Kim C. K., (2021). The ACC deaminase-producing plant growth-promoting bacteria: Influences of bacterial strains and ACC deaminase activities in plant tolerance to abiotic stress. Physiol. Plant. 173, 1992-2012. https://doi.org/10.1111/ppl.13545
Nascimento F. X., Rossi M. J., Soares C. R. F. S., McConkey B. J. and Glick B. R., (2014). New insights into 1-Aminocyclopropane-1-Carboxylate (ACC) deaminase phylogeny, evolution and ecological significance. PLoS One 9. https://doi.org/10.1371/journal.pone.0099168
Ose T., Fujino A., Yao M., Watanabe N., Honma M. and Tanaka I., (2003). Reaction intermediate structures of 1-aminocyclopropane-1-carboxylate deaminase: Insight into PLP-dependent cyclopropane ring-opening reaction. J. Biol. Chem., 278: 41069-41076. https://doi.org/10.1074/jbc.M305865200
Pramanik K. and Mandal N. C., (2022). Structural heterogeneity assessment among the isoforms of fungal 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase: a comparative in silico perspective. J. Genet. Eng. Biotechnol. 20: 18. https://doi.org/10.1186/s43141-021-00294-0
Sahoo R. K., Ansari M. W., Dangar T. K., Mohanty S. and Tuteja N., (2013). Phenotypic and molecular characterisation of efficient nitrogen-fixing Azotobacter strains from rice fields for crop improvement. Protoplasma 251: 511-523. https://doi.org/10.1007/s00709-013-0547-2
Saleem M., Arshad M., Hussain S. and Bhatti, A. S., 92007). Perspective of plant growth promoting rhizobacteria (PGPR) containing ACC deaminase in Stress Agriculture. J. Ind. Microbiol. Biotechnol., 34: 635-648. https://doi.org/10.1007/s10295-007-0240-6
Santhoshkumar R. and Yusuf A., (2020). In silico structural modeling and analysis of physicochemical properties of curcumin synthase (CURS1, CURS2, and CURS3) proteins of Curcuma longa. J. Genet. Eng. Biotechnol., 18. https://doi.org/10.1186/s43141-020-00041-x
Saravanakumar D. and Samiyappan R., (2007). ACC deaminase from Pseudomonas fluorescens mediated saline resistance in groundnut (Arachis hypogea) plants. J. Appl. Microbiol., 102: 1283-1292. https://doi.org/10.1111/j.1365-2672.2006.03179.x
Singh N. and Kashyap S., (2012). In silico identification and characterization of 1-Aminocyclopropane-1-Carboxylate deaminase from Phytophthora sojae. J. Mol. Model., 18: 4101-4111. https://doi.org/10.1007/s00894-012-1389-0
Singh S., Yadav S. K., Mishra P., Maurya R., Rana V., Yadav A. K., Singh A., Ram G. and Ramteke P. W., (2015). Comparative analysis of 1-aminocyclopropane-1-carboxylate (ACC) deaminase in selected plant growth promoting rhizobacteria (PGPR). J. Pure Appl. Microbiol., 9: 1587-1596.
Tao J. J., Chen H. W., Ma B., Zhang W. K., Chen S. Y. and Zhang J. S., (2015). The role of ethylene in plants under salinity stress. Front. Plant Sci., 6: 1-12. https://doi.org/10.3389/fpls.2015.01059
Thomas-Barry G., Martin C. St., Ramsubhag A., Eudoxie G. and Miller J. R., (2024). Multi-trait efficiency and interactivity of bacterial consortia used to enhance plant performance under water stress conditions. Microbiol. Res., 281: 127610. https://doi.org/https://doi.org/10.1016/j.micres.2024.127610
Vaseeharan B., Shanthi S., Chen J.C . and Espiñeira M., (2012). Molecular cloning, sequence analysis and expression of Fein-Penaeidin from the haemocytes of Indian white shrimp Fenneropenaeus indicus. Results Immunol., 2: 35-43. https://doi.org/10.1016/j.rinim.2012.02.001
Wang C., Knill E., Glick B. R. and Défago G., (2000). Effect of transferring 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase genes into Pseudomonas fluorescens strain CHA0 and its gacA derivative CHA96 on their growth-promoting and disease- suppressive capacities. Can. J. Microbiol., 46: 898-907. https://doi.org/10.1139/w00-071.
Waterhouse A., Bertoni M., Bienert S., Studer G., Tauriello G., Gumienny R., Heer F. T., De Beer T. A. P., Rempfer C., Bordoli L., Lepore R. and Schwede T., (2018). SWISS-MODEL: Homology modelling of protein structures and complexes. Nucleic Acids Res. 46, W296-W303. https://doi.org/10.1093/nar/gky427
