The MexAB-OprM efflux pump of Pseudomonas aeruginosa is central to multidrug resistance of this organism, which infects immunocompromised hospital patients. The MexA, MexB, and OprM subunits were assumed to function as the membrane fusion protein, the body of the transporter, and the outer membrane channel protein, respectively. For better understanding of this important xenobiotic transporter, we show the xray crystallographic structure of MexA at a resolution of 2.40 Å. The global MexA structure showed unforeseen new features with a spiral assembly of six and seven protomers that were joined together at one end by a pseudo 2-fold image. The protomer showed a new protein structure with a tandem arrangement consisting of at least three domains and presumably one more. The rod domain had a long hairpin of twisted coiled-coil that extended to one end. The second domain adjacent to the rod ␣-helical domain was globular and constructed by a cluster of eight short ␤-sheets. The third domain located distal to the ␣-helical rod was globular and composed of seven short ␤-sheets and one short ␣-helix. The 13-mer was shaped like a woven rattan cylinder with a large internal tubular space and widely opened flared ends. The 6-mer and 7-mer had a funnel-like structure consisting of a tubular rod at one side and a widely opened flared funnel top at the other side. Based on these results, we constructed a model of the MexAB-OprM pump assembly. The three pairs of MexA dimers interacted with the periplasmic ␣-barrel domain of OprM via the ␣-helical hairpin, the second domain interacted with both MexB and OprM at their contact site, and the third and disordered domains probably interacted with the distal domain of MexB. In this fashion, the MexA subunit connected MexB and OprM, indicating that MexA is the membrane bridge protein.

While biotechnological applications of arginine (Arg) as a solution additive that prevents protein aggregation are increasing, the molecular mechanism of its effects remains unclear. In this study, we investigated the Arg-lysozyme complex by high-resolution crystallographic analysis. Three Arg molecules were observed to be in close proximity to aromatic amino acid residues of the protein surface, and their occupancies gradually increased with increasing Arg concentration. These interactions were mediated by electrostatic, hydrophobic and cation-π interactions with the surface residues. The binding of Arg decreased the accessible surface area of aromatic residues by 40%, but increased that of charged residues by 10%. These changes might prevent intermolecular hydrophobic interactions by shielding hydrophobic regions of the lysozyme surface, resulting in an increase in protein solubility.

The crystal structure of the Pyrus pyrifolia pistil ribonuclease (S 3 -RNase) responsible for gametophytic selfincompatibility was determined at 1.5-Å resolution. It consists of eight helices and seven ␤-strands, and its folding topology is typical of RNase T 2 family enzymes. Based on a structural comparison of S 3 -RNase with RNase Rh, a fungal RNase T 2 family enzyme, the active site residues of S 3 -RNase assigned were His 33 and His 88 as catalysts and Glu 84 and Lys 87 as stabilizers of an intermediate in the transition state. Moreover, amino acid residues that constitute substrate binding sites of the two RNases could be superimposed geometrically. A hypervariable (HV) region that has an S-allele-specific sequence comprises a long loop and short ␣-helix. This region is far from the active site cleft, exposed on the molecule's surface, and positively charged. Four positively selected (PS) regions, in which the number of nonsynonymous substitutions exceeds that of synonymous ones, are located on either side of the active site cleft, and accessible to solvent. These structural features suggest that the HV or PS regions may interact with a pollen S-gene product(s) to recognize self and non-self pollen.Many flowering plants have a self-incompatibility system that recognizes the self or non-self between the pistil and pollen (tube) after pollination and suppresses growth of the self-pollen tube to prevent self-fertilization (1, 2). Gametophytic self-incompatibility (GSI) 1 is controlled genetically by a single locus (S-locus) with multiple alleles (1, 2). When a pollen grain lands on a stigma of the pistil, a process that discriminates as to whether an S-allele of the pollen matches one of the two Salleles of the pistil takes place. The pollen grain germinates on the stigma and grows into the style toward the embryo. If its S-allele matches one of the two S-alleles of the pistil, pollen tube growth is arrested in the style, and no fertilization takes place. In solanaceous, scrophulariaceous, and rosaceous plants that have GSI, the pistil glycoproteins that cosegregate with the S-alleles have been identified as ribonucleases of the RNase T 2 family (S-RNase) (3). McClure et al. (4) reported that pollen rRNA is degraded after self-pollination but not after crosspollination and suggested that GSI expression is mediated by degradation of the pollen rRNA of self-pollen tubes by S-RNase, leading to depletion of protein biosynthesis and the eventual arrest of tube growth. S-RNase has been confirmed necessary for GSI from results of gain-of-function and loss-of-function experiments on transgenic plants of solanaceous species (5, 6). Transgenic experiments also have shown that the RNase activity of S-RNase is necessary for GSI (7), which in petunia the carbohydrate moiety is not responsible for GSI (8), and that mutant S-RNase, which has lost RNase activity, acts as a dominant negative for GSI (9).Based on these findings, two models have been proposed to explain S-allele-specific inhibition of pollen tube growth; the...