PDBsum entry 1lax

Sugar binding protein

PDB id: 1lax

Contents

Protein chains

369 a.a. *

Ligands

GLC-GLC ×2

Waters ×813

* Residue conservation analysis

PDB id:

1lax

Name:

Sugar binding protein

Title:

Crystal structure of male31, a defective folding mutant of maltose- binding protein

Structure:

Maltose-binding protein mutant male31. Chain: a, c. Fragment: male31. Synonym: maltodextrin-binding protein mutant. Engineered: yes. Mutation: yes

Source:

Escherichia coli. Organism_taxid: 562. Gene: male. Expressed in: escherichia coli. Expression_system_taxid: 562.

Resolution:

1.85Å R-factor: 0.159 R-free: 0.202

Authors:

F.A.Saul,M.Mourez,B.Vulliez-Le Normand,N.Sassoon,G.A.Bentley, J.M.Betton

Key ref:

F.A.Saul et al. (2003). Crystal structure of a defective folding protein. Protein Sci, 12, 577-585. PubMed id: 12592028 DOI: 10.1110/ps.0235103

Date:

29-Mar-02 Release date: 04-Mar-03

Protein chains

P0AEX9  (MALE_ECOLI) -  Maltose/maltodextrin-binding periplasmic protein from Escherichia coli (strain K12)

Seq: 396 a.a.

Struc: 369 a.a.*

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.?

DOI no: 10.1110/ps.0235103

Protein Sci 12:577-585 (2003)

PubMed id: 12592028

Crystal structure of a defective folding protein.

F.A.Saul, M.Mourez, B.Vulliez-Le Normand, N.Sassoon, G.A.Bentley, J.M.Betton.

ABSTRACT

Maltose-binding protein (MBP or MalE) of Escherichia coli is the periplasmic receptor of the maltose transport system. MalE31, a defective folding mutant of MalE carrying sequence changes Gly 32-->Asp and Ile 33-->Pro, is either degraded or forms inclusion bodies following its export to the periplasmic compartment. We have shown previously that overexpression of FkpA, a heat-shock periplasmic peptidyl-prolyl isomerase with chaperone activity, suppresses MalE31 misfolding. Here, we have exploited this property to characterize the maltose transport activity of MalE31 in whole cells. MalE31 displays defective transport behavior, even though it retains maltose-binding activity comparable with that of the wild-type protein. Because the mutated residues are in a region on the surface of MalE not identified previously as important for maltose transport, we have solved the crystal structure of MalE31 in the maltose-bound state in order to characterize the effects of these changes. The structure was determined by molecular replacement methods and refined to 1.85 A resolution. The conformation of MalE31 closely resembles that of wild-type MalE, with very small displacements of the mutated residues located in the loop connecting the first alpha-helix to the first beta-strand. The structural and functional characterization provides experimental evidence that MalE31 can attain a wild-type folded conformation, and suggest that the mutated sites are probably involved in the interactions with the membrane components of the maltose transport system.

Selected figure(s)

Figure 1.
Figure 1. FkpA prevents MalE31 aggregation. Cells expressing chromosomally encoded MalE-wt or MalE31, transformed by pTrc99 or pTfkp, were grown in rich medium at 37°C for 3 h, and then fractionated from spheroplasts. Periplasmic (soluble) and membrane (insoluble) fractions, corresponding to 5 x 10^9 cells, were analyzed on SDS-polyacrylamide (12.5%) gel stained by Coomassie blue. (Lane 1), Strain PD28 ( malE) carrying pTrc99. (Lane 2) Strain MC4100 (malE) carrying pTrc99. (Lane 3) Strain JMB5 (malE31) carrying pTrc99. (Lane 4) JMB5 cells carrying pTfkp.

Figure 5.
Figure 5. Schematic view of the MalE31 structure. The mutated residues at positions 32 and 33 (top left) occur in an ß turn at an exposed position in the amino-terminal domain. Positions 13, 14, 38, and 63 located in the N-domain correspond to mutations affecting maltose transport (see text). The bound maltose substrate (middle) is located in a deep cleft between the amino- and carboxy-terminal domains.

The above figures are reprinted by permission from the Protein Society: Protein Sci (2003, 12, 577-585) copyright 2003.

Figures were selected by the author.