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In polarized epithelial cells, syntaxin 3 localizes to the apical plasma membrane and is involved in membrane fusion of apical trafficking pathways. We show that syntaxin 3 contains a necessary and sufficient apical targeting signal centered around a conserved FMDE motif. Mutation of any of three critical residues within this motif leads to loss of specific apical targeting. Modeling based on the known structure of syntaxin 1 revealed that these residues are exposed on the surface of a three-helix bundle. Syntaxin 3 targeting does not require binding to Munc18b. Instead, syntaxin 3 recruits Munc18b to the plasma membrane. Expression of mislocalized mutant syntaxin 3 in Madin-Darby canine kidney cells leads to basolateral mistargeting of apical membrane proteins, disturbance of tight junction formation, and loss of ability to form an organized polarized epithelium. These results indicate that SNARE proteins contribute to the overall specificity of membrane trafficking in vivo, and that the polarity of syntaxin 3 is essential for epithelial cell polarization.

The crystal structure of the helical linker domain and U-box domain of zebrafish CHIP (DrCHIP- HU) is presented. Results provide insight into conformational variability in the domain arrangement of CHIP and into U-box-mediated recruitment of UbcH5b for the ubiquitination of Hsp70 and Hsp90 substrates.

Many novel signal transduction domains are being identified in the wake of genome sequencing projects and improved sensitivity in homology-detection techniques. The functions of these domains are being discovered by hypothesis-driven experiments and structural genomics approaches. This article reviews the recent highlights of research on modular signaling domains, and the relative contributions and limitations of the various approaches being used.

The role of lipids in signal transduction with reference to phosphatidylinositol 3-phosphate directing endosomal localization of regulatory proteins is discused. Specific phosphate and hydroxyl group interactions distinguish phosphatidylinositol 3-phosphate from other lipids. The lipid binds to FYVE and PX domains found in intracellular membrane transport/signal transduction proteins and phagocyte proteins, respectively.

Results show that VHS domain containing human target of myb 1 proteins are localized to membranes. Crystallographic analysis of VHS domains indicates that helicies 2 and 4 are involved in the protein-protein interaction.

A study was conducted to show that chloride binding influences the Schiff base pK in the gecko P521 cone-type visual pigment. Purified photoreceptor outer segments were stored in buffer at -70 degrees C. Results indicated that the gecko P521 pigment supports an absorption maximum around 475 nm at pH 8.5. Findings also showed that the deprotonation of the chloride binding site is needed for the deprotonation of the Schiff base.

The structure of type II-beta phosphatidylinositol phosphate kinase (PIPK) shows a protein kinase ATP-binding core. It indicates that all phosphoinositide kinases are part of one superfamily. The flatness of the membrane-binding site, along with the shallowness of the substrate-binding pocket and its closeness to the membrane indicate that PIPKs phosphyorylate their substrates in situ in membranes. This allows the enzyme to avoid an energetic penalty for removing the substrate from its low-energy conformation within the membrane.

The effects of proton uptake and release in the photocycle of the bacteriorhodopsin mutant E204Q were analyzed by utilizing flash-induced absorbance, decay, and pH dependence studies on transformed E204Q mutant purple membrane. It was found that the photocycling time of mutant E204Q increased following substitution by glutamine altering the proton release uptake event from M to O intermediate. Furthermore, deprotonated negatively charged form of azides and weak acids catalyze deprotonation of D85 by acting as proton recipients.

Arg82 was replaced with Lys to analyse the role of Arg82 in the catalysis of proton transfer in bacteriorhodopsin, which has an intrinsic pK(sub a) of about 1.7 pH units lower than that of Arg. From the R82K mutant expressed in Halobacterium salinarium, it was found that the pK(sub a) of purple-to-blue transition at low temperature is about 3.6. At high pH in the blue-to-purple transition, the pK(sub a) was 8.0, the rate constant of dark adaption, k(sub da), is proportional to the fraction of blue membrane between pH = 2 and 10, and the efficiency of protein transfer in R82K under steady state is at least 40%.

Polyacrylamide gel electrophoresis, site-directed mutagenesis, sequence analysis and proton pumping measurements of the retinal binding site of bacteriorhodopsin (bR) reveal that Met118 cysteine (Cys) apoprotein cross-links with all-trans 4-bromoretinal, which contains bromine at the allylic position of the cyclohexyl ring. The position and cross-linking of Met118 Cys apoprotein indicate its presence inside the bR chromophore binding site near the C-4 cyclohexyl ring position. The chromophore does not cross-link with the wild-type or the 121 Cys pigments.

The CUE domain of the Vps9p sorting protein binds to monoubiquitin. The two binding surfaces of the CUE monomer are opposite to each other and cannot bind a ubiquitin molecule simultaneously. However, dimerization facilitates both sides to bind a monoubiquitin molecule.

The FYVE domains' interaction with the phosphatidylinositol three-phosphate membranes controls membrane trafficking and signalling pathways. Study of the FYVE domain structure revealed that this amino acid region is composed of two antiparallel beta sheets and an L helix stabilized by two Zn2+-binding clusters. It was also observed that the tip of the FYVE domain has basic and hydrophobic surfaces that promote nonspecific interactions with the phospholipid bilayer.