title: Boundary of the nucleotide-binding domain of Streptococcus ComA based on functional and structural analysis.
abstract: The ATP-binding cassette (ABC) transporter ComA is a key molecule essential for the first step of the quorum-sensing system of Streptococcus. The nucleotide binding domains (NBD) of Streptococcus mutans ComA with different N termini, NBD1 (amino acid residues 495-760), NBD2 (517-760), and NBD3 (528-760), were expressed, purified, and characterized. The shortest NBD3 corresponds to the region commonly defined as NBD in the database searches of ABC transporters. A kinetic analysis showed that the extra N-terminal region conferred a significantly higher ATP hydrolytic activity on the NBD at a neutral pH. Gel-filtration, X-ray crystallography, and mutational analyses suggest that at least four to five residues beyond the N-terminal boundary of NBD3 indeed participate in stabilizing the protein scaffold of the domain structure, thereby facilitating the ATP-dependent dimerization of NBD which is a prerequisite to the catalysis. These findings, together with the presence of a highly conserved glycine residue in this region, support the redefinition of the N-terminal boundary of the NBD of these types of ABC exporters.
authors.affiliation: Department of Biochemistry, Faculty of Nursing, Osaka Medical College, Osaka 569-8686, Japan. med002@art.osaka-med.ac.jp
authors.forename: Seiji
authors.initials: S
authors.lastname: Ishii
authors.forename: Takato
authors.initials: T
authors.lastname: Yano
authors.forename: Akihiro
authors.initials: A
authors.lastname: Okamoto
authors.forename: Takeshi
authors.initials: T
authors.lastname: Murakawa
authors.forename: Hideyuki
authors.initials: H
authors.lastname: Hayashi
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000409:Alanine / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D020134:Catalytic Domain; D002850:Chromatography, Gel; D002942:Circular Dichroism; D017124:Conserved Sequence; D018360:Crystallography, X-Ray; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D005998:Glycine / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009711:Nucleotides / Q000378:metabolism; D011487:Protein Conformation; D055503:Protein Multimerization; D013291:Streptococcus / Q000737:chemistry*
chemical_list: D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; C061262:ComA protein, Bacteria; D004268:DNA-Binding Proteins; D009711:Nucleotides; D000255:Adenosine Triphosphate; D000409:Alanine; D005998:Glycine