Folding of a mutant maltose-binding protein of Escherichia coli which forms inclusion bodies.

Abstract

The maltose-binding protein (MalE) of Escherichia coli is the periplasmic component of the transport system for malto-oligosaccharides. We have examined the characteristics of a Mal- mutant of malE corresponding to the double substitution Gly32 --> Asp/Ile33 --> Pro, MalE31, previously obtained by random mutagenesis. In vivo, the MalE31 precursor is efficiently processed, but the mature protein forms inclusion bodies in the periplasm. Furthermore, the accumulation of insoluble MalE31 is independent of its cellular localization; MalE31 lacking its signal sequence forms inclusion bodies in the cytoplasm. The native MalE31 protein can be purified by affinity chromatography from inclusion bodies after denaturation by 8 M urea. The renatured protein exhibits full maltose binding affinity (Kd= 9 x 10(-7) M), suggesting that its folded structure is similar to that of the wild-type protein. Unfolding/refolding experiments show that MalE31 is less stable (-5. 5 kcal/mol) than the wild-type protein (-9.5 kcal/mol) and that folding intermediates have a high tendency to form aggregates. In conclusion, the observed phenotype of cells expressing malE31 can be explained by a defective folding pathway of the protein. Study holds ProTherm entries: 5252, 5253 Extra Details: periplasm; cellular localization; signal sequence;,binding affinity; folded structure; folding pathway

Submission Details

ID: rSQC62sQ3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:29 p.m.

Version: 1

Publication Details
Betton JM;Hofnung M,J. Biol. Chem. (1996) Folding of a mutant maltose-binding protein of Escherichia coli which forms inclusion bodies. PMID:8626487
Additional Information

Study Summary

Number of data points 7
Proteins Maltose-binding periplasmic protein ; Maltose-binding periplasmic protein
Unique complexes 2
Assays/Quantities/Protocols Experimental Assay: Cm ; Experimental Assay: m ; Experimental Assay: dG_H2O ; Derived Quantity: ddG_H2O
Libraries Mutations for sequence KIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLEEKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKLIAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIAADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGETAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFLENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAFWYAVRTAVINAASGRQTVDEALKDAQTRITK

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P
 

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)


Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Maltose-binding periplasmic protein P0AEY0 MALE_ECO57
100.0 Maltose-binding periplasmic protein P0AEX9 MALE_ECOLI
94.3 Maltose-binding periplasmic protein P19576 MALE_SALTY
93.9 Maltose-binding periplasmic protein P18815 MALE_KLEAE
90.7 Maltose-binding periplasmic protein P41130 MALE_PHOLU