HIGH-RESOLUTION STRUCTURES OF THE LIGAND-BINDING DOMAIN OF THE WILD-TYPE BACTERIAL ASPARTATE RECEPTOR

The high-resolution structures of the wild-type periplasmic domain of the bacterial aspartate receptor have been determined in the absence a nd presence of bound aspartate to 1.85 and 2.2 Angstrom resolution, re spectively. As we reported earlier, in the refined structure of the co mplexed form of the crosslinked cysteine mutant receptor, the binding of the aspartate at the first site was mediated through four bridging water molecules while the second site showed an occupant electron dens ity that best fit a sulfate group, which was present in the crystalliz ation solution at high concentration. In the wild-type periplasmic dom ain structure two aspartate residues are bound per dimer, but with dif ferent occupancies. There exists a ''strong'' aspartate-binding site w hose binding is again mediated by four water molecules while the secon d site contains aspartate whose B-factor is about 10% higher, signifyi ng weaker binding. The interaction between the second, ''weaker'' aspa rtate with the three ligand-binding arginine side-chains is slightly d ifferent from the first site. The major difference is that there are t hree water molecules mediating the binding of aspartate at the second site, whereas in the first site there are four bridging water molecule s. The fact that aspartate-complexed crystals of the wild-type were gr own with a large excess aspartate while the cross-linked crystals were grown with equal molar aspartate may explain the difference in the st oichiometry observed. The conservation of the four bridging water mole cules in the strong aspartate site of both the cross-linked and wild-t ype periplasmic domain may reflect an important binding motif. The per iplasmic domain in the apo form is a symmetrical dimer, in which each of the subunits is equivalent, and the two aspartate binding sites are identical. Upon the binding of aspartate, the subunits are no longer symmetrical. The main difference between the aspartate-bound and unbou nd forms is in a small, rigid-body rotation between the subunits withi n a dimer. The rotation is similar in both direction and magnitude in the crosslinked and wild-type periplasmic domains. The presence of the second aspartate in the wild-type structure does not make any additio nal rotation compared to the single-site binding. The conservation of the small angular change in vitro suggests that the inter-subunit rota tion may have relevance to the understanding of the mechanism of trans membrane signal transduction in vivo. (C) 1996 Academic Press Limited