Maltose‐binding protein interacts simultaneously and asymmetrically with both subunits of the Tar chemoreceptor

Abstract

SummaryThe Tar chemotactic signal transducer of Escherichia coli mediates attractant responses to L-aspartate and to maltose. Aspartate binds across the subunit interface of the periplasmic receptor domain of a Tar homodimer. Maltose, in contrast, first binds to the periplasmic maltose-binding protein (MBP), which in its ligand-stabilized closed form then interacts with Tar. lntragenic complementation was used to determine the MBP-binding site on the Tar dimer. Mutations causing certain substitutions at residues Tyr-143, Asn-145, Gly-147, Tyr-149, and Phe-150 of Tar lead to severe defects in maltose chemotaxis, as do certain mutations affecting residues Arg-73, Met-76, Asp-77, and Ser-83. These two sets of mutations defined two complementation groups when the defective proteins were co-expressed at equal levels from compatible plasmids. We conclude that MBP contacts both subunits of the Tar dimer simultaneously and asymmetrically. Mutations affecting Met-75 could not be complemented, suggesting that this residue is important for association of MBP with each subunit of the Tar dimer. When the residues involved in interaction with MBP were mapped onto the crystal structure of the Tar periplasmic domain, they localized to a groove at the membrane-distal apex of the domain and also extended onto one shoulder of the apical region.

THE TWO-COMPONENT SIGNALING PATHWAY OF BACTERIAL CHEMOTAXIS: A Molecular View of Signal Transduction by Receptors, Kinases, and Adaptation Enzymes

“…In the case of the S. typhimurium aspartate receptor, a second aspartate molecule may still bind weakly, whereas the E. coli receptor exhibits true half-of-sites binding in which the second aspartate is virtually excluded . Similar halfof-sites binding has been observed for the E. coli serine chemoreceptor, and the binding proteins are also proposed to dock to the chemoreceptors in a half-of-sites fashion (Lin et al 1994, Stoddard & Koshland 1992, Gardina et al 1997. It follows that negative cooperativity is a general feature of attractant binding to the chemoreceptor family.…”

“…Genetic studies indicate that both domains of MBP dock to the extreme periplasmic end of the aspartate receptor, whereas an intersubunit complementation analysis suggests that the bound MBP molecule establishes simultaneous, asymmetric contacts on both receptor subunits , Gardina et al 1997. In particular, the receptor recognition elements appear to include loops at the ends of helices α1 and α2 in one subunit, and α4′ in the other subunit (prime denotes different subunits).…”

“…The serine-binding site likely possesses a very similar architecture (Jeffery & Koshland 1993, Yeh et al 1996, whereas the citrate-binding site differs at one coordinating position (T154A) but retains the three conserved arginine residues utilized in aspartate and serine binding (Yamamoto & Imae 1993). For large-molecule ligands, the docking of MBP probably represents typical binding protein-receptor interactions (Vyas et al 1991, Gardina et al 1997. Both small-and largemolecule attractants break the symmetry of the receptor dimer via negative cooperativity and asymmetric binding to the two receptor subunits.…”

Evidence for chemoreceptors with bimodular ligand-binding regions harboring two signal-binding sites

“…One mode consists of the direct recognition of chemoattractants like aspartate (7). The other mode is based on the interaction of the chemoattractant with a periplasmic binding protein, followed by the binding of the resulting complex to the Tar-LBR (8,9). Enterobacterial chemoreceptors contain an LBR composed of around 150 aa (10).…”

Crystal Structure of the 47-kDa Lipoprotein of Treponema pallidum Reveals a Novel Penicillin-binding Protein

“…Taken together, it is tempting to speculate that the enhanced PBP activity of Tp47 has been observable, at least in part, by virtue of the inhibitory action of zinc on the intrinsic ␤-lactamase activity of Tp47. Finally, the in vitro hydrolytic activity of wild-type Tp47 was inhibited by tazobactam, an inhibitor of class A ␤-lactamases (42,43), suggesting that competitive inhibition is active site directed. The apparent K i value for hydrolysis of penicillin by wild-type Tp47 was 26.95 Ϯ 0.35 nM.…”