3MBT: Structure Of Monomeric Blc From E. Coli

Citation:
Abstract
The first bacterial member of the lipocalin protein family, Blc, was identified in Escherichia coli as an outer membrane lipoprotein that is expressed under conditions of environmental stress. Previous crystallographic studies in space group P222 with two molecules per asymmetric unit, supported by static light-scattering experiments in solution, indicated that Blc may form a functional homodimer with lysophospholipid binding activity. Here, a new crystal structure of recombinant Blc in space group I422 with one molecule in the asymmetric unit is described. The crystal packing differs considerably from that observed previously, which was determined using an N-terminally extended version of Blc dubbed `Blc-X'. In particular, the characteristic large interface region that was previously described as being responsible for stable dimer formation is absent in the I422 crystal structure. Thus, the dimerization behaviour of Blc-X was most likely to be caused by the additional N-terminal peptide segment resulting from the cloning strategy employed. In contrast, we used a native-like N-terminus for Blc with just the lipid-anchored first Cys residue replaced by Ala. The fully monomeric status of this recombinant version of Blc in solution was confirmed by size-exclusion chromatography as well as analytical ultracentrifugation. Consequently, these data shed new light on the previously postulated lipid-binding mechanism and biological role of Blc. Beyond this, our findings illustrate that cloning artefacts, which frequently result from recombinant protein production for structural studies, must be considered with special caution when interpreting oligomerization and/or conformational effects.
PDB ID: 3MBTDownload
MMDB ID: 87164
PDB Deposition Date: 2010/3/26
Updated in MMDB: 2010/12
Experimental Method:
x-ray diffraction
Resolution: 2.6  Å
Source Organism:
Similar Structures:
Biological Unit for 3MBT: monomeric; determined by author and by software (PISA)
Molecular Components in 3MBT
Label Count Molecule
Protein (1 molecule)
1
Outer Membrane Lipoprotein BLC(Gene symbol: blc)
Molecule annotation
* Click molecule labels to explore molecular sequence information.

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