Proteomic approaches to identify substrates of the three Deg/HtrA proteases of the cyanobacterium Synechocystis sp. PCC 6803. Tam, L. X., Aigner, H., Timmerman, E., Gevaert, K., & Funk, C. Biochem J, 468(3):373–84, June, 2015. Edition: 2015/04/17![Proteomic approaches to identify substrates of the three Deg/HtrA proteases of the cyanobacterium Synechocystis sp. PCC 6803 [link]](https://bibbase.org/img/filetypes/link.svg "link")
Paper doi abstract bibtex The family of Deg/HtrA proteases plays an important role in quality control of cellular proteins in a wide range of organisms. In the genome of the cyanobacterium Synechocystis sp. PCC 6803, a model organism for photosynthetic research and renewable energy products, three Deg proteases are encoded, termed HhoA, HhoB and HtrA. In the present study, we compared wild-type (WT) Synechocystis cells with the single insertion mutants DeltahhoA, DeltahhoB and DeltahtrA. Protein expression of the remaining Deg/HtrA proteases was strongly affected in the single insertion mutants. Detailed proteomic studies using DIGE (difference gel electrophoresis) and N-terminal COFRADIC (N-terminal combined fractional diagonal chromatography) revealed that inactivation of a single Deg protease has similar impact on the proteomes of the three mutants; differences to WT were observed in enzymes involved in the major metabolic pathways. Changes in the amount of phosphate permease system Pst-1 were observed only in the insertion mutant DeltahhoB. N-terminal COFRADIC analyses on cell lysates of DeltahhoB confirmed changed amounts of many cell envelope proteins, including the phosphate permease systems, compared with WT. In vitro COFRADIC studies were performed to identify the specificity profiles of the recombinant proteases rHhoA, rHhoB or rHtrA added to the Synechocystis WT proteome. The combined in vivo and in vitro N-terminal COFRADIC datasets propose RbcS as a natural substrate for HhoA, PsbO for HhoB and HtrA and Pbp8 for HtrA. We therefore suggest that each Synechocystis Deg protease protects the cell through different, but connected mechanisms.
@article{tam_proteomic_2015,
title = {Proteomic approaches to identify substrates of the three {Deg}/{HtrA} proteases of the cyanobacterium {Synechocystis} sp. {PCC} 6803},
volume = {468},
issn = {1470-8728 (Electronic) 0264-6021 (Linking)},
url = {https://www.ncbi.nlm.nih.gov/pubmed/25877158},
doi = {10/f3pn99},
abstract = {The family of Deg/HtrA proteases plays an important role in quality control of cellular proteins in a wide range of organisms. In the genome of the cyanobacterium Synechocystis sp. PCC 6803, a model organism for photosynthetic research and renewable energy products, three Deg proteases are encoded, termed HhoA, HhoB and HtrA. In the present study, we compared wild-type (WT) Synechocystis cells with the single insertion mutants DeltahhoA, DeltahhoB and DeltahtrA. Protein expression of the remaining Deg/HtrA proteases was strongly affected in the single insertion mutants. Detailed proteomic studies using DIGE (difference gel electrophoresis) and N-terminal COFRADIC (N-terminal combined fractional diagonal chromatography) revealed that inactivation of a single Deg protease has similar impact on the proteomes of the three mutants; differences to WT were observed in enzymes involved in the major metabolic pathways. Changes in the amount of phosphate permease system Pst-1 were observed only in the insertion mutant DeltahhoB. N-terminal COFRADIC analyses on cell lysates of DeltahhoB confirmed changed amounts of many cell envelope proteins, including the phosphate permease systems, compared with WT. In vitro COFRADIC studies were performed to identify the specificity profiles of the recombinant proteases rHhoA, rHhoB or rHtrA added to the Synechocystis WT proteome. The combined in vivo and in vitro N-terminal COFRADIC datasets propose RbcS as a natural substrate for HhoA, PsbO for HhoB and HtrA and Pbp8 for HtrA. We therefore suggest that each Synechocystis Deg protease protects the cell through different, but connected mechanisms.},
language = {en},
number = {3},
urldate = {2021-06-07},
journal = {Biochem J},
author = {Tam, L. X. and Aigner, H. and Timmerman, E. and Gevaert, K. and Funk, C.},
month = jun,
year = {2015},
note = {Edition: 2015/04/17},
keywords = {Bacterial Proteins/genetics/*metabolism, Gene Deletion, Gene Expression Profiling, Gene Expression Regulation, Bacterial, Heat-Shock Proteins/genetics/*metabolism, Mutagenesis, Insertional, N-terminal combined fractional diagonal chromatography (COFRADIC), Periplasmic Proteins/genetics/*metabolism, Phosphate Transport Proteins/genetics/metabolism, Photosystem II Protein Complex/genetics/*metabolism, Protein Subunits/genetics/metabolism, Proteolysis, Proteomics/methods, Recombinant Proteins/genetics/metabolism, Ribulose-Bisphosphate Carboxylase/genetics/*metabolism, Serine Endopeptidases/genetics/*metabolism, Serine-Type D-Ala-D-Ala Carboxypeptidase/genetics/*metabolism, Substrate Specificity, Synechocystis 6803, Synechocystis/*enzymology/genetics/metabolism, Two-Dimensional Difference Gel Electrophoresis, difference gel electrophoresis (DIGE), expression, proteases},
pages = {373--84},
}
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In the present study, we compared wild-type (WT) Synechocystis cells with the single insertion mutants DeltahhoA, DeltahhoB and DeltahtrA. Protein expression of the remaining Deg/HtrA proteases was strongly affected in the single insertion mutants. Detailed proteomic studies using DIGE (difference gel electrophoresis) and N-terminal COFRADIC (N-terminal combined fractional diagonal chromatography) revealed that inactivation of a single Deg protease has similar impact on the proteomes of the three mutants; differences to WT were observed in enzymes involved in the major metabolic pathways. Changes in the amount of phosphate permease system Pst-1 were observed only in the insertion mutant DeltahhoB. N-terminal COFRADIC analyses on cell lysates of DeltahhoB confirmed changed amounts of many cell envelope proteins, including the phosphate permease systems, compared with WT. In vitro COFRADIC studies were performed to identify the specificity profiles of the recombinant proteases rHhoA, rHhoB or rHtrA added to the Synechocystis WT proteome. The combined in vivo and in vitro N-terminal COFRADIC datasets propose RbcS as a natural substrate for HhoA, PsbO for HhoB and HtrA and Pbp8 for HtrA. 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In the genome of the cyanobacterium Synechocystis sp. PCC 6803, a model organism for photosynthetic research and renewable energy products, three Deg proteases are encoded, termed HhoA, HhoB and HtrA. In the present study, we compared wild-type (WT) Synechocystis cells with the single insertion mutants DeltahhoA, DeltahhoB and DeltahtrA. Protein expression of the remaining Deg/HtrA proteases was strongly affected in the single insertion mutants. Detailed proteomic studies using DIGE (difference gel electrophoresis) and N-terminal COFRADIC (N-terminal combined fractional diagonal chromatography) revealed that inactivation of a single Deg protease has similar impact on the proteomes of the three mutants; differences to WT were observed in enzymes involved in the major metabolic pathways. Changes in the amount of phosphate permease system Pst-1 were observed only in the insertion mutant DeltahhoB. 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