Complete alanine scanning of the Escherichia coli RbsB ribose binding protein reveals residues important for chemoreceptor signaling and periplasmic abundance
| Autor: | Siham Beggah, Manuel Hernandez Gil, Vitali Maffenbeier, Artur Reimer, Manupriyam Dubey, Vladimir Sentchilo, Diogo Tavares, Jan Roelof van der Meer |
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| Jazyk: | angličtina |
| Rok vydání: | 2017 |
| Předmět: |
Models
Molecular 0301 basic medicine Ribose Science 030106 microbiology Mutant Molecular Conformation Plasma protein binding Biology Alanine/chemistry Alanine/metabolism Amino Acid Substitution Escherichia coli/genetics Escherichia coli/metabolism Escherichia coli Proteins/chemistry Escherichia coli Proteins/genetics Escherichia coli Proteins/metabolism Models Biological Molecular Imaging Mutation Periplasmic Binding Proteins/chemistry Periplasmic Binding Proteins/genetics Periplasmic Binding Proteins/metabolism Protein Binding Protein Transport Ribose/metabolism Signal Transduction medicine.disease_cause Article 03 medical and health sciences chemistry.chemical_compound Escherichia coli medicine Alanine Multidisciplinary Escherichia coli Proteins Binding protein Periplasmic space Alanine scanning Transport protein 030104 developmental biology chemistry Biochemistry Periplasmic Binding Proteins Medicine |
| Zdroj: | Scientific Reports, Vol 7, Iss 1, Pp 1-11 (2017) Scientific reports, vol. 7, no. 1, pp. 8245 Scientific Reports |
| ISSN: | 2045-2322 |
| Popis: | The Escherichia coli RbsB ribose binding protein has been used as a scaffold for predicting new ligand binding functions through in silico modeling, yet with limited success and reproducibility. In order to possibly improve the success of predictive modeling on RbsB, we study here the influence of individual residues on RbsB-mediated signaling in a near complete library of alanine-substituted RbsB mutants. Among a total of 232 tested mutants, we found 10 which no longer activated GFPmut2 reporter expression in E. coli from a ribose-RbsB hybrid receptor signaling chain, and 13 with significantly lower GFPmut2 induction than wild-type. Quantitative mass spectrometry abundance measurements of 25 mutants and wild-type RbsB in periplasmic space showed four categories of effects. Some (such as D89A) seem correctly produced and translocated but fail to be induced with ribose. Others (such as N190A) show lower induction probably as a result of less efficient production, folding and translocation. The third (such as N41A or K29A) have defects in both induction and abundance. The fourth category consists of semi-constitutive mutants with increased periplasmic abundance but maintenance of ribose induction. Our data show how RbsB modeling should include ligand-binding as well as folding, translocation and receptor binding. |
| Databáze: | OpenAIRE |
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