Deletion of a highly motional residue affects formation of the Michaelis complex for Escherichia coli dihydrofolate reductase

Analysis of the dihydrofolate reductase (DHFR) complex with folate by two-dimensional heteronuclear (1H-15N) nuclear magnetic relaxation revealed that isolated residues exhibit diverse backbone fluctuations on the nanosecond to picosecond time scale [Epstein, D. M., Benkovic, S. J., and Wright, P. E. (1995) Biochemistry 34, 11037-11048]. These dynamical features may be significant in forming the Michaelis complex. Of these residues, glycine 121 displays large-amplitude backbone motions on the nanosecond time scale. This amino acid, strictly conserved for prokaryotic DHFRs, is located at the center of the betaF-betaG loop. To investigate the catalytic importance of this residue, we report the effects of Gly121 deletion and glycine insertion into the modified betaF-betaG loop. Relative to wild type, deletion of Gly121 dramatically decreases the rate of hydride transfer 550-fold and the strength of cofactor binding 20-fold for NADPH and 7-fold for NADP+. Furthermore, DeltaG121 DHFR requires conformational changes dependent on the initial binary complex to attain the Michaelis complex poised for hydride transfer. Surprisingly, the insertion mutants displayed a significant decrease in both substrate and cofactor binding. The introduction of glycine into the modified betaF-betaG loop, however, generally eliminated conformational changes required by DeltaG121 DHFR to attain the Michaelis complex. Taken together, these results suggest that the catalytic role for the betaF-betaG loop includes formation of liganded complexes and proper orientation of substrate and cofactor. Through a transient interaction with the Met20 loop, alterations of the betaF-betaG loop can orchestrate proximal and distal effects on binding and catalysis that implicate a variety of enzyme conformations participating in the catalytic cycle.     

Biocenotic dynamics of liquid sewage in the process of its biological purification at aeration stations

A study was made of biological purification of sewage at the aeration stations on the quantitative composition of the main indicator microbes--of bacteria of the coliform group and of the fecal coliform bacilli, enterococci, Proteus, and also pathogenic enterobacteria. There was found a difference in the behaviour of different species of Proteus, i.e. reduction in the process of purification in the numbers of Pr. mirabilis, and a sharp elevation of Pr. morganii content. There was noted an insignificant amount of Pr. vulgaris both before and after the biological purification. It was found that dynamics of biocenosis was influenced by air temperature at the time of collection of the samples. A possibility of reproduction of coliform bacilli serving as one of the factors of autopurification of sewage during the biological purification was confirmed.     

Contested adoption cases: Grounds for conflict between psychology and the law.

Conflicting psychological testimony, statutory limitations, and judicial naivete with respect to child development concepts can often result in decisions in contested adoption cases that are not in the child's best interests. Illustrative case studies are presented, and 3 recommendations are made that might reduce this potentially harmful conflict between psychology and the law: (1) education of the legal profession, (2) more effective use of a child advocate system, and (3) legislative recognition of the concepts of children's rights and "psychological parenting." (36 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

MMPI overlap item scales for differentiating psychotics, neurotics, and nonpsychiatric groups.

To test the hypothesis that overlapping items of MMPI scales measure factors common to the criterion groups associated with the scales, 3 scales were constructed: Neurotic Overlap, Psychotic Overlap, and Maladjustment Overlap. These scales were validated against clinical psychotic and neurotic groups and 2 normal control groups. Results support the hypothesis and the validity of the 3 new scales. (2 ref) (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Computer techniques in the analysis of field-ion micrographs

The computer techniques at present used in the extraction and analysis of data from field-ion micrographs are described. These techniques are divided into three categories: simulation of field-ion images, the analysis of particle sizes and the determination of precipitate shapes and interface topography. The theory and techniques of each are described, together with examples of their use.     

Cellular expression and regulation of iron transport and storage proteins in genetic haemochromatosis

A series of ts mutations in the GSP1 gene of Saccharomyces cerevisiae was isolated by error-prone PCR. A total of 25 ts gsp1 strains was obtained. Each of these mutants showed between one and seven different amino acid alterations. In several of these ts gsp1 strains, the same amino acid residues in Gsp1p were repeatedly mutated, indicating that our screen for ts gsp1 mutations was saturating. All of the ts gsp1 strains isolated had a defect in nuclear protein import, but only 16 of the 25 ts gsp1 strains had a defect in mRNA export. Thus, Gsp1p is suggested to be directly involved in nuclear protein import, but not in mRNA export. Following release from alpha-factor arrest, 11 of the ts gsp1 mutants arrested in G1; the remainder did not show any specific cell-cycle arrest, at 37 degrees C, the nonpermissive temperature. While the mutants that are defective in both mRNA export and protein import have a tendency to arrest in G1, there was no clear correlation between the cell cycle phenotype and the defects in mRNA export and nuclear protein import. Based on this, we assume that Ran/Gsp1p GTPase regulates the cell cycle and the nucleus/cytosol exchange of macromolecules through interactions with effectors that were independent of each other, and are differentially affected by mutation.     

A comparison of guanosine-quartet inhibitory effects versus cytidine homopolymer inhibitory effects on rat neointimal formation

Biotinyl-oligosaccharides are a relatively new generation of saccharide probes that enable immobilization of desired oligosaccharides on streptavidin matrices for studies of carbohydrate-protein interactions. Here we describe the facile preparation of biotinyl-l-3-(2-naphthyl)-alanine hydrazide (BNAH) derivatives of oligosaccharides, containing a strong UV absorbing and fluorescent group, in which the ring of the reducing-end monosaccharide is nonreduced. We evaluate reactivities of immobilized BNAH- N -glycans with plant lectins that recognize aspects of the oligosaccharide core or outer-arms. We make some comparisons with 2-amino-6-amidobiotinyl-pyridine (BAP) derivatives obtained by reductive amination, and 6-(biotinyl)-aminocaproyl-hydrazide (BACH) derivatives which have a longer spacer-arm. N -Glycan-BNAH and-BAP derivatives have, overall, comparable reactivities with lectins which recognize N -glycan outer-arms or the trimannosyl core, but only BNAH and BACH derivatives are bound by lectins which recognize the non-reduced core. Moreover, with Pisum sativum agglutinin (PSA) which additionally requires the fucosyl- N- glycan-asparaginyl core for high affinity binding, the immobilized BNAH derivative (which is an alanine hydrazide beta-glycoside) can substitute for the natural beta-glycosylasparaginyl core, whereas the BACH derivative (aminocaproyl-hydrazide-beta-glycoside) is less effective. BNAH is a derivatization reagent of choice, therefore, for solid phase carbohydrate-binding experiments with immobilized N -glycans.