Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties
The role of glutamate as osmoprotector was investigated through the study of a mutation in its biosynthetic pathway. A glt::Tn917-lacZ-cat insertion mutant (N1) conferring glutamate auxotrophy and enhanced β-galactosidase expression on high-salt media was selected. Co-transformation experiments and...
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todo:paper_03438651_v47_n3_p208_Ruzal2023-10-03T15:26:15Z Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties Ruzal, S.M. Sanchez-Rivas, C. bacterial protein beta galactosidase glutamate synthase glutamic acid lysozyme article Bacillus subtilis bacterial mutation bacterial spore enzyme activity germination heat tolerance nonhuman priority journal radiosensitivity sporogenesis transposon Artificial Gene Fusion Bacillus subtilis Bacterial Proteins beta-Galactosidase DNA Transposable Elements Gene Expression Regulation, Bacterial Genes, Bacterial Genes, Reporter Glutamate Synthase Glutamic Acid Muramidase Mutagenesis, Insertional Osmotic Pressure Peptidoglycan Spores, Bacterial Transcription, Genetic Bacillus subtilis Bacteria (microorganisms) Felis catus insertion sequences Posibacteria transposons The role of glutamate as osmoprotector was investigated through the study of a mutation in its biosynthetic pathway. A glt::Tn917-lacZ-cat insertion mutant (N1) conferring glutamate auxotrophy and enhanced β-galactosidase expression on high-salt media was selected. Co-transformation experiments and PCR analysis allowed locating the insertion into the gltB gene corresponding to the small unit of the glutamate synthase (GOGAT). The N1 mutant strain presented a glutamate requirement for growth and a tenfold decrease in GOGAT activity. Transcriptional activity of GOGAT, measured as β-galactosidase from the transposon fusion, correlated with enzymatic activity; expression was enhanced at the stationary phase and in high-ionic-strength media. However, osmotolerance of cultures of N1 mutant were as wild-type (wt), at least in semi-rich medium. In contrast, sporulation was slightly reduced (75% of wt), and spores were less resistant to UV, heat, and osmolarity, properties linked to the content of small, acid-soluble proteins (SASP). The content of these proteins was, in fact, reduced, in particular the SASP-γ type. The peptidoglycan-cortex, however, was not impaired since spores maintained lysozyme resistance. Addition of glutamate during sporulation partially rescued spore resistance, but germination and outgrowth remained impaired. Deficiencies in germination and outgrowth were also observed with spores from a gltA mutant strain. Taken together, these results pointed to the importance of GOGAT activity during sporulation, in particular for the synthesis SASPs. Fil:Ruzal, S.M. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina. JOUR info:eu-repo/semantics/openAccess http://creativecommons.org/licenses/by/2.5/ar http://hdl.handle.net/20.500.12110/paper_03438651_v47_n3_p208_Ruzal |
| institution |
Universidad de Buenos Aires |
| institution_str |
I-28 |
| repository_str |
R-134 |
| collection |
Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) |
| topic |
bacterial protein beta galactosidase glutamate synthase glutamic acid lysozyme article Bacillus subtilis bacterial mutation bacterial spore enzyme activity germination heat tolerance nonhuman priority journal radiosensitivity sporogenesis transposon Artificial Gene Fusion Bacillus subtilis Bacterial Proteins beta-Galactosidase DNA Transposable Elements Gene Expression Regulation, Bacterial Genes, Bacterial Genes, Reporter Glutamate Synthase Glutamic Acid Muramidase Mutagenesis, Insertional Osmotic Pressure Peptidoglycan Spores, Bacterial Transcription, Genetic Bacillus subtilis Bacteria (microorganisms) Felis catus insertion sequences Posibacteria transposons |
| spellingShingle |
bacterial protein beta galactosidase glutamate synthase glutamic acid lysozyme article Bacillus subtilis bacterial mutation bacterial spore enzyme activity germination heat tolerance nonhuman priority journal radiosensitivity sporogenesis transposon Artificial Gene Fusion Bacillus subtilis Bacterial Proteins beta-Galactosidase DNA Transposable Elements Gene Expression Regulation, Bacterial Genes, Bacterial Genes, Reporter Glutamate Synthase Glutamic Acid Muramidase Mutagenesis, Insertional Osmotic Pressure Peptidoglycan Spores, Bacterial Transcription, Genetic Bacillus subtilis Bacteria (microorganisms) Felis catus insertion sequences Posibacteria transposons Ruzal, S.M. Sanchez-Rivas, C. Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties |
| topic_facet |
bacterial protein beta galactosidase glutamate synthase glutamic acid lysozyme article Bacillus subtilis bacterial mutation bacterial spore enzyme activity germination heat tolerance nonhuman priority journal radiosensitivity sporogenesis transposon Artificial Gene Fusion Bacillus subtilis Bacterial Proteins beta-Galactosidase DNA Transposable Elements Gene Expression Regulation, Bacterial Genes, Bacterial Genes, Reporter Glutamate Synthase Glutamic Acid Muramidase Mutagenesis, Insertional Osmotic Pressure Peptidoglycan Spores, Bacterial Transcription, Genetic Bacillus subtilis Bacteria (microorganisms) Felis catus insertion sequences Posibacteria transposons |
| description |
The role of glutamate as osmoprotector was investigated through the study of a mutation in its biosynthetic pathway. A glt::Tn917-lacZ-cat insertion mutant (N1) conferring glutamate auxotrophy and enhanced β-galactosidase expression on high-salt media was selected. Co-transformation experiments and PCR analysis allowed locating the insertion into the gltB gene corresponding to the small unit of the glutamate synthase (GOGAT). The N1 mutant strain presented a glutamate requirement for growth and a tenfold decrease in GOGAT activity. Transcriptional activity of GOGAT, measured as β-galactosidase from the transposon fusion, correlated with enzymatic activity; expression was enhanced at the stationary phase and in high-ionic-strength media. However, osmotolerance of cultures of N1 mutant were as wild-type (wt), at least in semi-rich medium. In contrast, sporulation was slightly reduced (75% of wt), and spores were less resistant to UV, heat, and osmolarity, properties linked to the content of small, acid-soluble proteins (SASP). The content of these proteins was, in fact, reduced, in particular the SASP-γ type. The peptidoglycan-cortex, however, was not impaired since spores maintained lysozyme resistance. Addition of glutamate during sporulation partially rescued spore resistance, but germination and outgrowth remained impaired. Deficiencies in germination and outgrowth were also observed with spores from a gltA mutant strain. Taken together, these results pointed to the importance of GOGAT activity during sporulation, in particular for the synthesis SASPs. |
| format |
JOUR |
| author |
Ruzal, S.M. Sanchez-Rivas, C. |
| author_facet |
Ruzal, S.M. Sanchez-Rivas, C. |
| author_sort |
Ruzal, S.M. |
| title |
Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties |
| title_short |
Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties |
| title_full |
Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties |
| title_fullStr |
Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties |
| title_full_unstemmed |
Effect of glutamate synthase (GOGAT) activity on Bacillus subtilis spore properties |
| title_sort |
effect of glutamate synthase (gogat) activity on bacillus subtilis spore properties |
| url |
http://hdl.handle.net/20.500.12110/paper_03438651_v47_n3_p208_Ruzal |
| work_keys_str_mv |
AT ruzalsm effectofglutamatesynthasegogatactivityonbacillussubtilissporeproperties AT sanchezrivasc effectofglutamatesynthasegogatactivityonbacillussubtilissporeproperties |
| _version_ |
1782023545483689984 |