A general rule for the relationship between hydrophobic effect and conformational stability of a protein: stability and structure of a series of hydrophobic mutants of human lysozyme

Previous research has shown that head direction (HD) cells in both the anterior dorsal thalamus (ADN) and the postsubiculum (PoS) in rats discharge in relation to familiar, visual landmarks in the environment. This study assessed whether PoS and ADN HD cells would be similarly responsive to nonvisual or unfamiliar environmental cues. After visual input was eliminated by blindfolding the rats, HD cells maintained direction-specific discharge, but their preferred firing directions became less stable. In addition, rotations of the behavioral apparatus indicated that some nonvisual cues (presumably tactile, olfactory, or both) exerted above chance stimulus control over a cell's preferred firing direction. However, a prominent auditory cue was not effective in exerting stimulus control over a cell's preferred direction. HD cell activity also was assessed after rotation of a novel visual cue exposed to the rat for 1, 3, or 8 min. An 8-min exposure was enough time for a novel visual cue to gain control over a cell's preferred direction, whereas an exposure of 1 or 3 min led to control in only about half the sessions. These latter results indicate that HD cells rely on a rapid learning mechanism to develop associations with landmark cues.     

Anomalous mole fraction effect, electrostatics, and binding in ionic channels

Ionic channels bathed in mixed solutions of two permeant electrolytes often conduct less current than channels bathed in pure solutions of either. For many years, this anomalous mole fraction effect (AMFE) has been thought to occur only in single-file pores containing two or more ions at a time. Most thinking about channels incorporates this view. We show here that the AMFE arises naturally, as an electrostatic consequence of localized ion specific binding, if the average current through a channel is described by a theory (Poisson-Nernst-Planck, PNP) that computes the average electric field from the average concentration of charges in and near the channel. The theory contains only those ion-ion interactions mediated by the mean field, and it does not enforce single filing. The AMFE is predicted by PNP over a wide range of mean concentrations of ions in the channel; for example, it is predicted when (on the average) less, or much less, than one ion is found in the channel's pore. In this treatment, the AMFE arises, in large measure, from a depletion layer produced near a region of ion-specific binding. The small excess concentration of ions in the binding region repels all nearby ions of like charge, thereby creating a depletion layer. The overall conductance of the channel arises in effect from resistors in series, one from the binding region, one from the depletion zone, and one from the unbinding region. The highest value resistor (which occurs in the depletion zone) limits the overall series conductance. Here the AMFE is not the result of single filing or multiple occupancy, and so previous views of permeation need to be revised: the presence of an AMFE does not imply that ions permeate single file through a multiply occupied pore.     

Conserved sequence preference in DNA binding among recombination proteins: an effect of ssDNA secondary structure

Repetitive sequences have been proposed to be recombinogenic elements in eukaryotic chromosomes. We tested whether dinucleotide repeats sequences are preferential sites for recombination because of their high affinity for recombination enzymes. We compared the kinetics of the binding of the scRad51, hsRad51 and ecRecA proteins to oligonucleotides with repeats of dinucleotides GT, CA, CT, GA, GC or AT. Since secondary structures in single-stranded DNA (ssDNA) act as a barrier to complete binding we measured whether these oligonucleotides are able to form stable secondary structures. We show that the preferential binding of recombination proteins is conserved among the three proteins and is influenced mainly by secondary structures in ssDNA.     

Anchor structure of staphylococcal surface proteins. III. Role of the FemA, FemB, and FemX factors in anchoring surface proteins to the bacterial cell wall

Surface proteins of Staphylococcus aureus are covalently linked to the bacterial cell wall by a mechanism requiring a COOH-terminal sorting signal with a conserved LPXTG motif. Cleavage between the threonine and the glycine of the LPXTG motif liberates the carboxyl of threonine to form an amide bond with the pentaglycyl cross-bridge in the staphylococcal peptidoglycan. Here, we asked whether altered peptidoglycan cross-bridges interfere with the sorting reaction and investigated surface protein anchoring in staphylococcal fem mutants. S. aureus strains carrying mutations in the femA, femB, femAB, or the femAX genes synthesize altered cross-bridges, and each of these strains displayed decreased sorting activity. Characterization of cell wall anchor structures purified from the fem mutants revealed that surface proteins were linked to cross-bridges containing one, three, or five glycyl residues, but not to the epsilon-amino of lysyl in muropeptides without glycine. When tested in a femAB strain synthesizing cross-bridges with mono-, tri-, and pentaglycyl as well as tetraglycyl-monoseryl, surface proteins were found anchored mostly to the five-residue cross-bridges (pentaglycyl or tetraglycyl-monoseryl). Thus, although wild-type peptidoglycan appears to be the preferred substrate for the sorting reaction, altered cell wall cross-bridges can be linked to the COOH-terminal end of surface proteins.     

Main-chain conformational features at different conformations of the side-chains in proteins

An analysis of the known protein structures has shown that the main-chain torsion angles, phi and psi of a residue can be affected by the side-chain torsion angle, chi1. The (chi1, psi) plot of all residues (except Gly, Ala and Pro) show six distinct regions where points are concentrated-although some of these regions are nearly absent in specific cases. The mean of these clusters can show a shift along the psi axis by as much as 30 degrees as chi1 is changed from around 180 to -60 to 60 degrees. Because of the lesser steric constraint points are more diffused along the psi axis when chi1 is approximately -60 degrees. Although points are more spread out along the phi axis in the (chi1, phi) plot, the dependence of phi on chi1 shows up in a shortened phi range (by about 30 degrees) when chi1 is around -60 degrees, and a distinct tendency of clustering of points into two regions when chi1 is approximately equal to 60 degrees, especially for the aromatic residues. Based on the dependence of the backbone conformation on its side-chain the 17 amino acids can be grouped into five classes: (i) aliphatic residues branched at the Cbeta position (although Thr is atypical), (ii) Leu (branched at the Cgamma position), (iii) aromatic residues (Trp can show some deviations), (iv) short polar residues (Asp and Asn), and (v) the remaining linear-chain residues, mainly polar. Ser and Thr have the highest inclination to occur with two different orientations of the side-chain that can be located through crystallography. Such residues exhibiting two chi1 angles have their phi and psi angles in a region that is common to the Ramachandran plots at the two different chi1 angles. The dependence of phi and psi angles on chi1 can be used to understand the helical propensities of some residues. Moreover, the average phi, psi values in the alpha-helices vary with the side-chain conformation.     

Role, personality, and social structure in the organizational setting.

An analysis of the interaction between the psychological factors of the individual functioning within the environmental context of an organizational structure (hospital, business, prison, school, etc.) and the resultant "role" played. 36 refs. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Effect of natural ligands on the structural properties and conformational stability of proteins

The effect of natural ligands on the structural properties and conformational stability of proteins is reviewed. It is shown that the range of possible structural transformations induced in a protein molecule by ligand release is very wide and virtually does not depend on the nature of the protein or that of the ligands. Ligand-free forms of protein are classified from the viewpoint of structural property changes of a protein molecule.     

Role of the ninaC proteins in photoreceptor cell structure: ultrastructure of ninaC deletion mutants and binding to actin filaments

The ninaC proteins are found in Drosophila photoreceptor cells. Their primary sequences suggest they are kinase/myosin chimeras, but their myosin head-like domain is the most divergent amongst all the myosin-like proteins described to date. To investigate possible roles of the ninaC proteins in cell structure, we examined the ultrastructure of the photoreceptor cells in various ninaC mutants, and tested the ability of the proteins to interact with actin filaments in a myosin-like manner. In flies lacking the larger ninaC protein, p174, an ultrastructural phenotype was evident before eclosion. The axial actin cytoskeleton of the rhabdomeral microvilli appeared either fragmented or as an isolated structure, without linkage to the microvillar membrane. Deletion of the myosin head-like domain or the calmodulin-binding domain of p174 resulted in a similar abnormal cytoskeleton. Breakdown of the rhabdomeres followed, although at different rates depending on the deletion. Lack of the smaller protein, p132, per se did not result in photoreceptor degeneration, but in older flies there was an abnormal accumulation of multivesicular bodies. Moreover, the presence of p132 retarded the degeneration that occurs in the absence of p174, even though the p132 remained outside the rhabdomere. Biochemical studies showed that both ninaC proteins bind actin filaments and cosediment with actin filaments in an ATP-sensitive manner. These results outline structural roles for the ninaC proteins, and are consistent with the notion, suggested by their amino acid sequences, that the proteins are actin-based mechanoenzymes.     

Comparison of ultra- and microfiltration in the presence and absence of secondary flow with polysaccharides, proteins, and yeast suspensions

The purpose of this research was to show that controlled centrifugal instabilities-Dean vortices-produced by solutions and suspensions from typical biotechnology applications flowing through curved tubes can be used to reduce concentration polarization and/or fouling in pressure-driven ultrafiltration (UF) and microfiltration (MF) processes. Experiments were conducted to (i) evaluate the ultrafiltration performance of hollow fiber membranes in linear and helical configurations with dextran (low fouling) and bovine serum albumin (high fouling) solutions and (ii) compare the performance of linear and helical coiled UF hollow fiber modules with that of similar MF modules using baker's and beer yeast (Saccharomyces cerevisiae) suspensions as feed. Both constant transmembrane pressure (TMP) and constant permeation flux (J) experiments were utilized here. The membrane material was polyether sulfone. For the ultrafiltration experiments, the helical module performed consistently better than the linear module with dextran T500 and BSA solutions, resulting in performance improvements (helical versus linear) from 20 to 200% and up to 85%, respectively. For the comparative experiments between UF and MF, the helical module again performed better than the linear module for low concentration baker's yeast suspensions (0.5-1% dry wt). At constant TMP, the flux improvements for UF were 30-120%, while at constant J, the capacity or loading was 4.5 times higher for the UF as compared to the MF membrane. At high beer yeast concentrations (5.1-6.8% dry wt), although flux improvements were not observed between the linear and helical modules for UF, the UF fluxes were 72% higher than that obtained with MF. Also, for MF, with the same high beer yeast concentrations, the helical module exhibited 30-90% higher fluxes than that obtained with the linear module. At constant flux (117-137 L m-2 h-1) and intermediate baker's yeast concentrations (0.65-2.7% dry wt), 10-20 times the capacity was obtained for the helical over the linear module. Yeast cells were the dominant foulant. For constant UF flux (70 L m-2 h-1) experiments at high beer yeast concentrations ((4.3-7.7) x 10(7) cells/mL or 5.1-6.8% dry wt), the capacity (loading) for the helical module was 10 times that of the linear module. Again, the yeast cells were the dominant foulant. A new mass-transfer correlation for ultrafiltration of dextran T500 solutions for laminar flow in a helical hollow fiber module was obtained, viz. Sh = 0.173Re0.55Sc0.33(a/Rc)0.07.     

Mucin genes and the proteins they encode: structure, diversity, and regulation

Mucins are the structural components of the mucus gels that protect the respiratory, gastrointestinal, and reproductive tracts. These polydisperse glycoproteins (250,000 to 20,000,000 D) are approximately 80% carbohydrate on a mass basis and have a high intrinsic viscosity due to their large size and extreme hydrophilicity. Mucin oligosaccharides, the structures responsible for this hydrophilicity, are heterogeneous in size and structure but are chiefly O-linked, i.e., they initiate from N-acetylgalactosamine residues attached to threonine and serine residues of the polypeptide backbone. Our understanding of the structure of mucins has advanced rapidly in the last few years with the isolation and sequencing of cDNA clones that encode mucin polypeptide backbones. All currently well-characterized mucins have been found to contain extended arrays of tandemly repeated peptides rich in potential O-glycosylation sites. Less is known about the unique sequences that flank the tandem repeat arrays of secretory mucins, but currently available information indicates that these flanking regions contain cysteine-rich stretches that participate in mucin oligomer formation. Thus, secretory mucins appear to consist of oligomers containing heavily glycosylated domains flanked by unique sequences required for polymerization. Progress has also been made in characterizing the genes that encode mucins. At least four human mucin genes are known at present, although many others may remain to be discovered. Moreover, much work remains before we gain an understanding of the mechanisms involved in the expression of mucin genes and their tissue-specific regulation.     

Resonance assignments, secondary structure and topology of leukaemia inhibitory factor in solution

We evaluated a protocol involving two types of choice presentations for assessing leisure choice-making skills of seven older adults with severe disabilities. Initially when presented with pairs of objects representing choices, choice making was validated through demonstration of an object preference. A more complex choice-presentation format was then employed, involving pictures to represent choices. If the preference identified with objects was not demonstrated using pictures, a replication of the object format occurred to ensure changes in choice making using pictures was not due to a preference change. Five participants demonstrated choice-making skills using objects and two demonstrated choices using pictures. These results reflect the importance of assessing choice-making skills prior to presenting choice opportunities. Suggestions for future research focus on expanding the assessment protocol to include a wider array of choice-making skills and training staff to provide choices in a format commensurate with an individual's skill level.     

The pH dependence of hydrogen-deuterium exchange in trp repressor: the exchange rate of amide protons in proteins reflects tertiary interactions, not only secondary structure

The pH dependence of amide proton exchange rates have been measured for trp-repressor. One class of protons exchanges too fast to be measured in these experiments. Among the protons that have measurable hydrogen-deuterium exchange rates, two additional classes may be distinguished. The second class of protons are in elements of secondary structure that are mostly on the surface of the protein, and exchange linearly with increasing base concentration (log kex versus pH). The third class of amide protons is characterized by much higher protection against exchange at higher pH. These protons are located in the core of the protein, in helices B and C. The exchange rate in the core region does not increase linearly with pH, but rather goes through a minimum around pH 6. The mechanism of exchange for the slowly exchanging core protons is interpreted in terms of the two-process model of Hilton and Woodward (1979, Biochemistry 18:5834-5841), i.e., exchange through both a local mechanism that does not require unfolding of the protein, and a mechanism involving global unfolding of the protein. The increase in exchange rates at low pH is attributed to a partial unfolding of the repressor. It is concluded that the formation of secondary structure alone is insufficient to account for the high protection factors seen in the core of native proteins at higher pH, and that tertiary interactions are essential to stabilize the structure.     

Prediction of homology and divergence in the secondary structure of polypeptides

A quantitative procedure is described for the comparison of secondary structure of homologous proteins. Standard predictive methods are used to generate probability profiles from pairs of homologous amino acid sequences; correlation coefficients (R) are then computed between each pair of amino acids for alpha-helix (R alpha), extended structure (R beta), turn (R(t)), and coil (R(c)). R values are >0.2 for correctly aligned homologous sequences. Unrelated or incorrectly aligned sequences give R values near zero. Lack of correlation for a segment of otherwise well-correlated sequences is used to identify structural divergence, which is then evaluated graphically by using difference profiles. A combination of these techniques correctly predicts secondary structural differences between melittin or beta-endorphin and their respective synthetic analogs. The method is potentially useful to describe evolutionary changes in protein secondary structure as well as in the design of peptide analogs.