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A Wierzbicki

Publications and source records attributed to A Wierzbicki.

At least 37 records · Page 2Linked to original sources
PubMed1995

Scanning electron microscopy and molecular modeling of inhibition of calcium oxalate monohydrate crystal growth by citrate and phosphocitrate.

Binding of citrate and phosphocitrate to calcium oxalate monohydrate crystals has been studied using scanning electron microscopy (SEM) and molecular modeling. Phosphocitrate structure has been resolved using low temperature X-ray analysis and ab initio computational methods. The (-1 0 1) crystal surface of calcium oxalate monohydrate is involved in binding of citrate and phosphocitrate, as shown by SEM and molecular modeling. Citrate and phosphocitrate conformations and binding energies to (-1 0 1) faces have been obtained and compared to binding to another set of calcium-rich planes (0 1 0). Difference in inhibitory properties of these compounds has been attributed to better coordination of functional groups of phosphocitrate with calcium ions in (-1 0 1). Relevance of this study to design of new calcium oxalate monohydrate inhibitors is discussed.

Calcium Oxalate↗
PubMed1994

Atomic force microscopy and molecular modeling of protein and peptide binding to calcite.

Oyster shell protein and polyaspartate bound to calcite have been visualized at the atomic and molecular levels by atomic force microscopy. The identities of potential binding sites have been suggested from atomic force microscopy (AFM) images and have been evaluated by molecular modeling. Energies and conformations of binding to (110) and (110) prism faces, (001) basal calcium planes, and (104) cleavage planes are considered. The interaction with the basal plane is strongest and is essentially irreversible. Binding to (110) prism surfaces is also energetically favored and selective for orientations parallel or perpendicular to the c-axis. Binding to (110) faces is significantly weaker and orientation nonspecific. If carboxyl groups of the protein or peptide replace select carbonate ions of the (110) face, the binding energy increases significantly, favoring binding in the parallel direction. Binding to (104) cleavage surfaces is weak and probably reversible. Specific alignment of oyster shell protein molecules on calcite surfaces is shown by AFM, and the relevance to the binding model is discussed.

Animals↗
PubMed1993

Cloning of higher plant omega-3 fatty acid desaturases.

Arabidopsis thaliana T-DNA transformants were screened for mutations affecting seed fatty acid composition. A mutant line was found with reduced levels of linolenic acid (18:3) due to a T-DNA insertion. Genomic DNA flanking the T-DNA insertion was used to obtain an Arabidopsis cDNA that encodes a polypeptide identified as a microsomal omega-3 fatty acid desaturase by its complementation of the mutation. Analysis of lipid content in transgenic tissues demonstrated that this enzyme is limiting for 18:3 production in Arabidopsis seeds and carrot hairy roots. This cDNA was used to isolate a related Arabidopsis cDNA, whose mRNA is accumulated to a much higher level in leaf tissue relative to root tissue. This related cDNA encodes a protein that is a homolog of the microsomal desaturase but has an N-terminal extension deduced to be a transit peptide, and its gene maps to a position consistent with that of the Arabidopsis fad D locus, which controls plastid omega-3 desaturation. These Arabidopsis cDNAs were used as hybridization probes to isolate cDNAs encoding homologous proteins from developing seeds of soybean and rapeseed. The high degree of sequence similarity between these sequences suggests that the omega-3 desaturases use a common enzyme mechanism.

Algorithms↗
PubMed1990

Alteration of the lipid composition of rat testicular plasma membranes by dietary (n-3) fatty acids changes the responsiveness of Leydig cells and testosterone synthesis.

Experiments were conducted to assess whether changing dietary fat composition altered phospholipid composition of rat testicular plasma membranes in a manner that altered receptor-mediated action of luteinizing hormone (LH)/human chorionic gonadotropin (hCG). Weanling rats were fed diets that provided high or low cholesterol intakes and that were enriched with linseed oil, fish oil or beef tallow for 4 wk. Feeding diets high in (n-3) fatty acids decreased plasma and testicular plasma membrane 20:4(n-6) content. A marked reduction of the 22:5(n-6) content and an increase in the 22:6(n-3) content of testicular plasma membrane was found only in animals fed fish oil. A decrease in binding capacity of the gonadotropin (LH/hCG) receptor in the plasma membrane, with no change in receptor affinity, was observed for animals fed either linseed oil or fish oil diets. Dietary treatments that raised plasma membrane cholesterol content and the cholesterol to phospholipid ratio in the membrane were associated with increased binding capacity of the gonadotropin receptor. Feeding diets high in 18:3(n-3) vs. those high in fish oil altered receptor-mediated adenylate cyclase activity in a manner that depended on the level of dietary cholesterol. Feeding diets high in cholesterol or fish oil increased basal and LH-stimulated testosterone synthesis relative to that in animals fed the low cholesterol diet containing linseed oil. It is concluded that changing the fat composition of the diet alters the phospholipid composition of rat testicular plasma membranes and that this change in composition influences membrane-mediated unmasking of gonadotropin receptor-mediated action in testicular tissue.

Adenylyl Cyclases↗
PubMed1990

Alterations in dietary fatty acid composition alter rat brush border membrane phospholipid fatty acid composition.

Female Wistar rats were fed for 8 weeks a high polyunsaturated fatty acid diet (P) enriched with 18:2w6 and 18:3w3, or a high saturated fatty acid diet (S) enriched with 16:0, 18:0 and 18:1w9. There was a similar jejunal BBM fatty acid profile in phosphatidylcholine (PC) and phosphatidylethanolamine (PE) composition in animals fed P. However, feeding S altered the fatty acid composition of BBM phospholipids: for PC, feeding S increased total monounsaturated fatty acids (e.g. 18:1(9)) and reduced total polyunsaturated fatty acids, especially the total w6 fatty acids. For PE, feeding S increased only the total monounsaturated fatty acids. The ratio of monounsaturated/saturated fatty acids was increased for PC and PE. Fatty acid changes were more pronounced in the jejunal than ileal BBM. It is concluded that alterations in dietary fatty acid saturation: (a) influences intestinal BBM phospholipid fatty acid composition; (b) these changes are quantitatively and qualitatively different for PC and PE; and (c) changes are greater in the proximal than in the distal intestine. It is proposed that the previously reported diet-associated changes in active and passive intestinal transport are due at least in part to these alterations in the fatty acid composition in brush border membrane phospholipids.

Animals↗
PubMed1989

Fish oil prevents change in arachidonic acid and cholesterol content in rat caused by dietary cholesterol.

Rats were fed diets high in either saturated fat (beef tallow) or alpha-linolenic acid (linseed oil) or eicosapentaenoic and docosahexaenoic acids (fish oil) with or without 2% cholesterol supplementation. Consumption of linseed oil and fish oil diets for 28 days lowered arachidonic acid content of plasma, liver and heart phospholipids. Addition of 2% cholesterol to diets containing beef tallow or linseed oil lowered 20:4 omega 6 levels but failed to reduce 20:4 omega 6 levels when fed in combination with fish oil. Feeding omega 3 fatty acids lowered plasma cholesterol levels. Addition of 2% cholesterol to the beef tallow or linseed oil diet increased plasma cholesterol concentrations but not when fish oil was fed. Feeding the fish oil diet reduced the cholesterol content of liver, whereas feeding the linseed oil diet did not. Dietary cholesterol supplementation elevated the cholesterol concentration in liver in the order: linseed oil greater than beef tallow greater than fish oil (8.6-, 5.5-, 2.6-fold, respectively). Feeding fish oil and cholesterol apparently reduced 20:4 omega 6 levels in plasma and tissue lipids. Fish oil accentuates the 20:4 omega 6 lowering effect of dietary cholesterol and appears to prevent accumulation of cholesterol in plasma and tissue lipids under a high dietary load of cholesterol.

Animals↗
PubMed1988

Properties of a mutant Cre protein that alters the topological linkage of recombination products.

The bacteriophage P1 Cre-loxP site-specific recombination system consists of two components: the Cre recombinase protein and the loxP DNA sequence where recombination takes place. We report here on the analysis of a mutation in the cre structural gene that produces a mutant protein with altered recombination properties. The mutant protein, Cre111, carries out recombination at a much slower rate than the wild-type Cre protein. To determine why the reaction is slow, we have examined a number of activities associated with Cre-mediated recombination. Our results indicate that the binding of Cre111 to the loxP site is comparable to wild-type Cre. Furthermore, the rate at which Cre111 resolves Holliday structures, an intermediate in this recombination reaction, is also comparable to wild-type Cre. Thus, DNA binding and resolution of the intermediate are not affected, suggesting that either synapsis, the process of bringing two lox sites together, or the first strand exchange event, could be affected in the mutant protein. The types of DNA products formed following recombination of a supercoiled substrate can reflect mechanisms of synapsis. Wild-type Cre generates mainly topologically unlinked and unknotted circular products. This suggests that the wild-type protein brings two lox sites together in a way that excludes the entanglement of supercoils present in the substrate DNA. In contrast, when Cre111 recombines a supercoiled molecule it generates many complicated catenanes and knotted DNA products. Presumably, the supercoils present in the DNA substrate are being trapped in the reaction products.(ABSTRACT TRUNCATED AT 250 WORDS)

Coliphages↗
PubMed1988

Differential effects of dietary linoleic and alpha-linolenic acid on lipid metabolism in rat tissues.

Comparative effects of feeding dietary linoleic (safflower oil) and alpha-linolenic (linseed oil) acids on the cholesterol content and fatty acid composition of plasma, liver, heart and epididymal fat pads of rats were examined. Animals fed hydrogenated beef tallow were used as isocaloric controls. Plasma cholesterol concentration was lower and the cholesterol level in liver increased in animals fed the safflower oil diet. Feeding the linseed oil diet was more effective in lowering plasma cholesterol content and did not result in cholesterol accumulation in the liver. The cholesterol concentration in heart and the epididymal fat pad was not affected by the type of dietary fatty acid fed. Arachidonic acid content of plasma lipids was significantly elevated in animals fed the safflower oil diet and remained unchanged by feeding the linseed oil diet, when compared with the isocaloric control animals fed hydrogenated beef tallow. Arachidonic acid content of liver and heart lipids was lower in animals fed diets containing safflower oil or linseed oil. Replacement of 50% of the safflower oil in the diet with linseed oil increased alpha-linolenic, docosapentaenoic and docosahexaenoic acids in plasma, liver, heart and epididymal fat pad lipids. These results suggest that dietary 18:2 omega 6 shifts cholesterol from plasma to liver pools followed by redistribution of 20:4 omega 6 from tissue to plasma pools. This redistribution pattern was not apparent when 18:3 omega 3 was included in the diet.

Adipose Tissue↗
PubMed1988

Influence of genotype on the phospholipid fatty acid composition of splenic T and B lymphocytes in MRL/MpJ-lpr/lpr mice.

The MRL/MpJ-lpr/lpr mice manifest a T cell proliferative and autoimmune disorder. Similar changes occur much later in the life of MRL/MpJ-+/+ mice. MRL/MpJ-lpr/lpr (lpr/lpr) and MRL/MpJ-+/+ (+/+) mice were fed for six weeks nutritionally adequate semipurified diets containing 20% (w/w) fat, but differing in linoleic acid content. The phospholipid fatty acid composition of T and B cells was found to be dependent on genetic background of mice and level of linoleic acid in the diet. Changes in the levels of specific fatty acids like 16:0, 18:2 omega 6, 22:5 omega 3 and 22:6 omega 3 in some of the phospholipid components were observed in the MRL/MpJ-lpr/lpr strain in both the B and T cell types as compared with their normal +/+ counterpart strain. T cells of lpr/lpr mice exhibited significantly higher levels of 20:4 omega 6 than did T cells of other strain. High levels of dietary linoleic acid significantly increased incorporation of 18:2 omega 6 in T and B cells, while the effect on other fatty acids of the two types of cells varied with the phospholipid classes and fatty acids when compared with the low linoleic acid fed-group. Differences observed in the phospholipid fatty acid composition of the T and B cells of the congenic mice might contribute to differences in rate of progression of age-related changes suggesting that the autoimmune disorder might be mitigated by dietary manipulation.

Animals↗
PubMed1988

Regional differences in lipid composition and incorporation of saturated and unsaturated fatty acids into microsomal membranes of rat small intestine.

Incorporation of [1-14C]palmitic (16:0) and [1-14C]linoleic (18:2 omega 6) acids into microsomal membranes of proximal (jejunum) and distal (ileum) regions of rat small intestine was investigated, and the lipid composition, including fatty acid profiles of membrane phospholipids, was determined. Jejunal microsomes contained significantly higher amounts of total phospholipids, phosphatidylcholine, and phosphatidylinositol, and lower amounts of cholesterol and sphingomyelin when compared with ileal microsomes. Jejunal microsomal phospholipids contained higher levels of stearic (18:0), 18:2 omega 6, and eicosapentaenoic (20:5 omega 3) acids followed by reduced levels of oleic (18:1 omega 9), arachidonic (20:4 omega 6), and docosahexaenoic (22:6 omega 3) acids when compared with those from the ileum, except for phosphatidylinositol where no significant difference between 20:4 omega 6 content of each site was observed. In both jejunal and ileal microsomes, incorporation of [1-14C]18:2 omega 6 was significantly higher than that of [1-14C]16:0. Incorporation of both [1-14C]16:0 and [1-14C]18:2 omega 6 was significantly higher in jejunal microsomal lipid fractions (phospholipids, diacylglycerols, triacylglycerols) when compared with the ileal microsomal fraction. These data suggest that (1) jejunal and ileal microsomal membranes differ from each other in terms of lipid composition and lipid synthesis, (2) site variations in the specificity of acyltransferases for different fatty acids exist, and (3) higher delta 9-, delta 6-, delta 5-, and delta 4-desaturase activities exist in ileal compared with jejunal enterocytes.

Animals↗
PubMed1988

Modulation of rat testes lipid composition by hormones: effect of PRL and hCG.

The effect of prolactin (PRL) and human chorionic gonadotropin (hCG) administration for 7 days on the composition and function of rat testicular plasma membrane was investigated. Refractory state in Leydig cells desensitized by hCG decreased the binding capacity for 125I-labeled hCG and also luteinizing hormone (LH)-induced adenosine 3',5'-cyclic monophosphate (cAMP) and testosterone production. In testicular membranes of hCG-treated animals, a depletion of cholesterol and an increase in total phospholipid content was observed after gonadotropin injection, thereby decreasing the cholesterol-to-phospholipid ratio. Injection of high doses of PRL had no effect on the binding capacity or affinity of the LH-hCG receptor but decreased the response of Leydig cells to LH in terms of cAMP and testosterone synthesis. PRL also increased total and esterified cholesterol and decreased free cholesterol and membrane phospholipid content. The fatty acid composition of testicular lipids was significantly and selectively influenced by both hormonal treatments. These observations suggest that metabolism of cholesterol and long-chain polyunsaturated fatty acids in testicular tissue is affected by chorionic gonadotropin and PRL and may provide the mechanism for regulating steroidogenic functions.

Animals↗
PubMed1987

A mutational analysis of the bacteriophage P1 recombinase Cre.

Bacteriophage P1 encodes a 38,600 Mr site-specific recombinase, Cre, that is responsible for reciprocal recombination between sites on the P1 DNA called loxP. Using in vitro mutagenesis 67 cre mutants representing a total of 37 unique changes have been characterized. The mutations result in a wide variety of phenotypes as judged by the varying ability of each mutant Cre protein to excise a lacZ gene located between two loxP sites in vivo. Although the mutations are found throughout the entire cre gene, almost half are located near the carboxyl terminus of the protein, suggesting a region critical for recombinase function. DNA binding assays using partially purified mutant proteins indicate that mutations in two widely separated regions of the protein each result in loss of heparin-resistant complexes between Cre and a loxP site. These results suggest that Cre may contain two separate domains, both of which are involved in binding to loxP.

Amino Acid Sequence↗
PubMed1987

Isolation and characterization of intermediates in site-specific recombination.

Cre, the site-specific recombinase from bacteriophage P1, catalyzes a recombination reaction between specific DNA sequences designated as lox sites. The breakage and rejoining of partners during this recombination process must be highly concerted because it has not been possible to detect intermediates of the reaction with wild-type Cre. Several mutant Cre proteins have been isolated that produce significant amounts of a possible intermediate product of the recombination reaction. The product has been identified as a Holliday structure in which one set of the DNA strands of the recombining partners has been exchanged. Wild-type Cre protein is capable of acting on this structure to form recombinant products, which is consistent with this being an intermediate in the recombination reaction. Characterization of the Holliday structure indicated that one set of strands in the recombining partners was always exchanged preferentially before the other set. In addition, it has been found that certain Cre mutants that are unable to carry out recombination in vitro are able to resolve the intermediate. This suggests that these mutants are defective in a step in the reaction that precedes the formation of the Holliday intermediate.

Coliphages↗
PubMed1986

The role of the loxP spacer region in P1 site-specific recombination.

The lox-Cre site-specific recombination system of bacteriophage P1 is comprised of a site on the DNA where recombination occurs called loxP, and a protein, Cre, which mediates the reaction. The loxP site is 34 base pairs (bp) in length and consists of two 13 bp inverted repeats separated by an 8 bp spacer region. Previously it has been shown that the cleavage and strand exchange of recombining loxP sites occurs within this spacer region. We report here an analysis of various base substitution mutations within the spacer region of loxP, and conclude the following: Homology is a requirement for efficient recombination between recombining loxP sites. There is at least one position within the spacer where a base change drastically reduces recombination even when there is homology between the two recombining loxP sites. When two loxP sites containing symmetric spacer regions undergo Cre-mediated recombination in vitro, the DNA between the sites undergoes both excision and inversion with equal frequency.

Bacterial Proteins↗