Independent folding of the domains in the hydrophilic subunit IIABman of the mannose transporter of Escherichia coli.


Abstract

The active form of the hydrophilic subunit (IIABman) of the mannose transporter of Escherichia coli is a homodimer of two 35-kDa subunits. Each subunit consists of two distinct domains, IIA and IIB, which can be separated by limited trypsin digestion. Separation of tryptic fragments yields monomers of IIB and dimers of IIA, which are active and stable. To test whether the domains fold as independent units, the effects of guanidine hydrochloride (GuHCl) and temperature on the structural stability of the intact IIABman were compared with those of the isolated fragments. Equilibrium GuHCl-induced reversible unfolding, measured by circular dichroism and tryptophan fluorescence, showed a biphasic transition for intact IIABman and monophasic transitions for each isolated fragment. The midpoint transitions of the isolated IIB and IIA fragments (at 1.0 and 2.3 M GuHCl) coincide with the first and second transitions of intact IIABman. Analytical ultracentrifugation studies suggested that dissociation precedes the unfolding of IIA. Thermal unfolding of IIABman, monitored by differential scanning calorimetry, showed two well-separated transitions near 52 and 95 degrees C which corresponded to the midpoint transitions of the isolated IIB and IIA fragments. The combined results demonstrate an independent stepwise unfolding of the domains in IIABman as well as the absence of stabilizing interdomain interactions. The lack of interdomain interactions suggests an unrestricted domain motion. This may play an important role in the phosphoryl transfer reaction which is catalyzed by the binding of IIABman to a phosphoryl carrier protein HPr (via the IIA domain) and to the transmembrane subunits of the mannose transporter (via the IIB domain). Study holds ProTherm entries: 4804, 4805, 4806 Extra Details:

Submission Details

ID: 9BVxb3Vf

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:27 p.m.

Version: 1

Publication Details
Marković-Housley Z;Cooper A;Lustig A;Flükiger K;Stolz B;Erni B,Biochemistry (1994) Independent folding of the domains in the hydrophilic subunit IIABman of the mannose transporter of Escherichia coli. PMID:8086415
Additional Information

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 PDB Entries

Structure ID Release Date Resolution Structure Title
2JZN 2008-02-19 Solution NMR structure of the productive complex between IIAMannose and IIBMannose of the mannose transporter of the E. coli phosphotransferase system
2JZO 2008-02-19 Solution NMR structure of the non-productive complex between IIAMannose and IIBMannose of the mannose transporter of the E. coli phosphotransferase system
1VRC 2005-04-19 Complex of enzyme IIAmannose and the histidine-containing phosphocarrier protein HPr from escherichia coli nmr, restrained regularized mean structure
2JZH 2008-02-19 structure of IIB domain of the mannose transporter of E. coli
1VSQ 2008-02-19 Solution NMR structure of the productive complex between IIAMannose and IIBMannose of the mannose transporter of the E. coli phosphotransferase system
1PDO 1996-08-01 1.7 PHOSPHOENOLPYRUVATE-DEPENDENT PHOSPHOTRANSFERASE SYSTEM

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 PTS system mannose-specific EIIAB component P69800 PTNAB_SHIFL
100.0 PTS system mannose-specific EIIAB component P69797 PTNAB_ECOLI
100.0 PTS system mannose-specific EIIAB component P69798 PTNAB_ECOL6
100.0 PTS system mannose-specific EIIAB component P69799 PTNAB_ECO57