Structural basis for the altered activity of Gly794 variants of Escherichia coli beta-galactosidase

The open-closed conformational switch in the active site of Escherichia coli beta-galactosidase was studied by X-ray crystallography and enzyme kinetics. Replacement of Gly794 by alanine causes the apoenzyme to adopt the closed rather than the open conformation. Binding of the competitive inhibitor isopropyl thio-beta-D-galactoside (IPTG) requires the mutant enzyme to adopt its less favored open conformation, weakening affinity relative to wild type. In contrast, transition-state inhibitors bind to the enzyme in the closed conformation, which is favored for the mutant, and display increased affinity relative to wild type. Changes in affinity suggest that the free energy difference between the closed and open forms is 1-2 kcal/mol. By favoring the closed conformation, the substitution moves the resting state of the enzyme along the reaction coordinate relative to the native enzyme and destabilizes the ground state relative to the first transition state. The result is that the rate constant for galactosylation is increased but degalactosylation is slower. The covalent intermediate may be better stabilized than the second transition state. The substitution also results in better binding of glucose to both the free and the galactosylated enzyme. However, transgalactosylation with glucose to produce allolactose (the inducer of the lac operon) is slower with the mutant than with the native enzyme. This suggests either that the glucose is misaligned for the reaction or that the galactosylated enzyme with glucose bound is stabilized relative to the transition state for transgalactosylation.     

Cortical cytoskeletal ring in prophase II leads to correction of abnormalities of the first meiotic division and to meiotic restitution of pollen mother cell nucleus

The deviation of prophase cytoskeletal ring formation was determined during meiotic division in 50% of pollen mother cells (PMCs) in maize haploid No 1498 (Zea mays). At prophase in both meiotic divisions the cytoskeletal ring is formed in cortical region of cytoplasm instead of perinuclear. Sometimes formation of both perinuclear and cortical rings is observed in the same cell. It has been shown that in multinucleate PMCs the cortical ring leads to the consolidation of chromosomes into common spindle and to meiotic restitution.     

Bouquet arrest and its consequences for asynaptic meiosis in cereals--haploids and wide hybrids F1

Arrest of chromosomes at bouquet configuration at zygothene leads to meiotic restitution in some haploids and allohaploids in cereals: univalents are transported preferentially to one spindle pole in bipolar spindle because of their oriented position in the beginning of prometaphase.     

Consolidation of cytoskeleton during plant spindle formation. II. "Fused spindles"

Three mechanisms of fused spindle formation in meiosis of Solanacea have been described: 1) approach of daughter nuclei at prophase II; 2) fusion of perinuclear cytoskeleton systems at prophase II; 3) approach and fusion of prometaphase chaotic figures at prometaphase II. The process of fusion spindle formation appears to be complex and including several steps.     

Consolidation of cytoskeleton during plant spindle formation. I. Abnormalities of spindle integrity in meiosis

The process of formation of multiple spindles in mononucleate pollen mother cells during meiosis in general wide hybrids F1 and haploids is described. It is assumed that the abnormality may be caused by aberration of a special process at late prometaphase providing for the spindle consolidation.     

Latitudinal Adh allozymic variation and ethanol tolerance in Indian populations of Drosophila melanogaster

Ten Indian geographical populations of D. melanogaster were assayed electrophoretically for Adh genic variation. The Indian geographical populations of D. melanogaster revealed significant clinal variation (3 % for 1 d? latitude) at Adh locus and Adh^F allelic frequency correlated significantly with increase in latitude. It was suggested that the abundance of secondary alcohols in the southern Indian tropical and humid environment might exert selective pressure favouring higher frequency of Adh^S allele. Patterns of ethanol utilization as well as ethanol tolerance were analyzed in larval and adult individuals of six geographical populations of D. melanogaster. Latitudinal variation in ethanol tolerance was observed in D. melanogaster populations from India. The parallel occurrence of latitudinal variation it Adh locus as well as ethanol tolerance in Indian geographical populations of D. melanogaster could be maintained by balancing natural selection varying spatially along the north-south axis of the Indian sub-continent.     

Microbial expression profiles in the rhizosphere of two maize lines differing in N use efficiency

Plant and Soil - In Ontario, Canada, acreage of red clover (Trifolium pratense L.) intercropped with winter wheat (Triticum aestivum L. em. Thell) has declined, despite well-documented soil and...     

Stereoselective effects of chiral clofibric acid analogs on rat peroxisome proliferator-activated receptor α (rPPARα) activation and peroxisomal fatty acid β-oxidation

Enantiomers of a series of substituted analogs of 2-(4-chloronhenoxy)-acetic acid (CPAA) were synthesized and used to examine the influence of steric and structural parameters on peroxisome proliferation. The effects of these compounds were studied on the activation of the peroxisome proliferator-activated receptor α (PPARα) in CV-1 cells using an in vitro co-transfection assay. Selected sets of isomers were tested for their ability to increase peroxisomal fatty acyl-CoA oxidase (ACO) activity in H4IIEC3 (rat Reuber hepatoma) cells. Of the series of 2-substituted analogs studied, the isomers of the n-propyl and phenyl derivatives of CPAA showed a high degree of stereoselectivity [(S)-isomer ≫ (R)-isomer]. In general, the potency of the compound to activate the receptor increased with the size of the 2-alkyl substituent. Among the 4-chlorobenzyloxy- and 4-(4′-chlorophenyl)benzyloxy- analogs studied, 2-[4-(4′-chlorophenyl)-benzyloxy]-propanoic acid exhibited a high degree of stereoselectivity in both the biological systems studied [(R) ≫ (S)]. The congeners of 2-methyl substituted CPAA showed a reverse stereoselectivity [(R) > (S)] as compared to the other 2-substituted analogs [(S) > (R)]. Our results indicate that (1) both structural and steric characteristics of CPAA analogs play an important role in the activation of rPPARα and on stimulation of peroxisomal ACO activities, and (2) clofibric acid and analogs exert their peroxisome proliferative effects by interaction with a specific site on a protein. The enantiomers of the 2-n-propyl and the 2-phenyl CPAA analogs may be useful as mechanistic probes in elucidating the nature of this binding site. Chirality 9:37–47, 1997. © 1997 Wiley-Liss, Inc.     

Processes providing the phragmoplast centrifugal movement in cereal meiosis

A group of new abnormalities in meiotic division in the mother cells of cereal pollen that affect the processes of the centrifugal movement of a phragmoplast in successive cytokinesis is presented for the first time. These phenotypes represent new information on the formation and operation of motile phragmoplast and confirm our model of centrifugal movement as B-anaphase modification.     

Estrogen-related Receptor �� Is a Key Regulator of Muscle Mitochondrial Activity and Oxidative Capacity

Estrogen-related receptor γ (ERRγ) regulates the perinatal switch to oxidative metabolism in the myocardium. We wanted to understand the significance of induction of ERRγ expression in skeletal muscle by exercise. Muscle-specific VP16ERRγ transgenic mice demonstrated an increase in exercise capacity, mitochondrial enzyme activity, and enlarged mitochondria despite lower muscle weights. Furthermore, peak oxidative capacity was higher in the transgenics as compared with control littermates. In contrast, mice lacking one copy of ERRγ exhibited decreased exercise capacity and muscle mitochondrial function. Interestingly, we observed that increased ERRγ in muscle generates a gene expression profile that closely overlays that of red oxidative fiber-type muscle. We further demonstrated that a small molecule agonist of ERRβ/γ can increase mitochondrial function in mouse myotubes. Our data indicate that ERRγ plays an important role in causing a shift toward slow twitch muscle type and, concomitantly, a greater capacity for endurance exercise. Thus, the activation of this nuclear receptor provides a potential node for therapeutic intervention for diseases such as obesity, which is associated with reduced oxidative metabolism and a lower type I fiber content in skeletal muscle.