The compatible-solute-binding protein OpuAC from Bacillus subtilis: ligand binding, site-directed mutagenesis, and crystallographic studies.
In the soil bacterium Bacillus subtilis, five transport systems work in concert to mediate the import of various compatible solutes that counteract the deleterious effects of increases in the osmolarity of the environment. Among these five systems, the ABC transporter OpuA, which catalyzes the import of glycine betaine and proline betaine, has been studied in detail in the past. Here, we demonstrate that OpuA is capable of importing the sulfobetaine dimethylsulfonioacetate (DMSA). Since OpuA is a classic ABC importer that relies on a substrate-binding protein priming the transporter with specificity and selectivity, we analyzed the OpuA-binding protein OpuAC by structural and mutational means with respect to DMSA binding. The determined crystal structure of OpuAC in complex with DMSA at a 2.8-A resolution and a detailed mutational analysis of these residues revealed a hierarchy within the amino acids participating in substrate binding. This finding is different from those for other binding proteins that recognize compatible solutes. Furthermore, important principles that enable OpuAC to specifically bind various compatible solutes were uncovered.
Smits
Sander H J
SH
Institute of Biochemistry, Heinrich Heine University Düesseldorf, Universitätsstr. 1, 40225 Düsseldorf, Germany.
Höing
Marina
M
Lecher
Justin
J
Jebbar
Mohamed
M
Schmitt
Lutz
L
Bremer
Erhard
E
D019943:Amino Acid Substitution / Q000235:genetics; D001412:Bacillus subtilis / Q000737:chemistry* / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D008074:Lipoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D015394:Molecular Structure; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D013379:Substrate Specificity; D013452:Sulfonium Compounds / Q000378:metabolism*
D001426:Bacterial Proteins; D008074:Lipoproteins; C108613:OpuAC protein, Bacillus subtilis; D013452:Sulfonium Compounds; C121105:dimethylsulfonioacetate