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Biomedical subjects

R G Wallace

Publications and source records attributed to R G Wallace.

At least 37 records · Page 2Linked to original sources
PubMed1985

Hydrophobic interactions of human milk lipase.

The hydrolysis of a series of n-alkyl esters of 4-nitrobenzoic acid, and of isopropyl 4-nitrobenzoate, 4'-nitrophenyl 4-nitrobenzoate, and 4-nitrobenzoyl 1-monoglycerol, catalyzed by human milk lipase in the absence and presence of cholate stimulation, has been measured at pH 7.3, 37.5 degrees C. It has been shown that the enzyme possesses a specific alkyl binding site which is hydrophobic in nature and wide enough to accommodate two fatty acid chains lying side by side or a phenyl ring lying flat. The hydrophobic nature of this site is affected by bile salt stimulation of the enzyme. Hydrophobicity parameters have been calculated for hydrocarbon chains lying in the acyl and alkyl binding sites of human milk lipase. A mechanism is suggested for the role of bile salts in stimulating the enzyme in its activity against water soluble esters and water soluble triacylglycerols.

Chemical Phenomena↗
PubMed1985

Human milk lipase substrates: electronic role.

The hydrolysis of a series of 4'-nitrophenyl 4-substituted benzoates, catalyzed by human milk lipase in the absence and presence of cholate stimulation, has been measured at pH 7.3, 37.5 degrees C. There is no evidence for an acyl site electronic interaction in the rate determining step of the esterolytic reaction. Substrate inhibition, arising from restricted rotation of the aryl residues about the ester linkage in the esterolytic site, is exhibited by these substrates.

Acylation↗
PubMed1985

Esterase acyl binding site of human milk lipase.

The hydrolysis of a series of 4-nitrophenyl alkanoates and of 4-nitrophenylbenzoate and -methylbenzoate, catalysed by human milk lipase in the absence and presence of cholate stimulation, has been measured at pH 7.3, 37.5 degrees C. It has been shown that the enzyme possesses a specific esterase acyl binding site that is hydrophobic in nature and wide enough to accommodate one fatty acid chain, but not two. This binding site is unaffected by bile salt stimulation of the enzyme.

Bile Acids and Salts↗
PubMed1984

Studies in bile salt solutions. The effect of pH on the cholate and taurocholate stimulation of human milk lipase catalyzed hydrolysis of p-nitrophenylacetate.

The pseudo-first-order rate constants of hydrolysis of p-nitrophenylacetate, catalyzed by human milk lipase, have been measured in solutions of 0.01 mol dm-3 Bistris(2-[bis(2-hydroxyethyl)amino]-2-(hydroxymethyl)-propane-1,3 -diol) buffer at 310.5 K, containing a range of concentrations of sodium taurocholate and sodium cholate, at pH 8.00 and of sodium cholate at pH 6.5. The effect of pH on the activity of the enzyme has been investigated and the stimulation factors of taurocholate and cholate ions and of cholic acid have been calculated to be equal to 5.3, 3.7 and 10.7, respectively. The essential residues for catalytic activity of the enzyme have ionization constants equal to 6.45-6.46 for pK1 and 8.33-8.40 for pK2. The former value is attributed to the presence of a histidine imidazolium group but the identity of the residue leading to pK2 is not proven.

Catalysis↗
PubMed1984

Studies in bile salt solutions. Deoxycholate stimulation of human milk lipase.

Stimulation of human milk lipase by deoxycholate and its taurine and glycine conjugates was demonstrated by measuring the esterolysis reaction of 4-nitrophenylacetate. The steroidal surfactants did not bind strongly to the polar substrate but they did bind effectively to a hydrophobic site on the enzyme and these bile salt-enzyme complexes were effective catalysts. These results are compared with those for stimulation of the enzyme by cholate surfactants and it has been demonstrated that the absence of a 7 alpha-OH substituent on the steroid nucleus does not prevent stimulation of either the esterolytic or lipolytic activity of the enzyme.

Deoxycholic Acid↗
PubMed1981

Mechanism of araC autoregulation and the domains of two overlapping promoters, Pc and PBAD, in the L-arabinose regulatory region of Escherichia coli.

The DNA-protein contact sites in the ara regulatory region, which contains the promoters for araBAD and araC, have been determined for araC protein, the cyclic AMP-binding protein, and RNA polymerase, by using the methylation protection and DNase I protection methods. The functional significance of binding was assessed by correlating the state of occupancy of these sites with promoter activity in transcription initiation. Our results suggest that the basis for araC autoregulation is that araC protein, in either its activator (P2) or repressor (P1) form, acts as a repressor for araC, by binding to the RNA polymerase attachment site at the araC promoter. We also found that the araC and araBAD promoters share a common site of positive control by the cyclic AMP-binding protein, located 90 bases from the araBAD and 60 bases from the araC transcriptional start points. A model for the mechanism of regulation of araBAD and araC expression by the catabolite gene-activator protein, P1, and Pe is proposed. An earlier model proposed by Ogden et al. [Ogden S., Haggerty, D., Stoner, C. M., Kolodrubetz, D. & Schleif, R. (1980) Proc. Natl. Acad. Sci, USA 77, 3346-3350] is discussed in the light of the data presented in this paper.

AraC Transcription Factor↗
PubMed1980

The araC gene of Escherichia coli: transcriptional and translational start-points and complete nucleotide sequence.

The nucleotide sequence of the Escherichia coli araC gene and flanking regions have been determined from a series of overlapping fragments using the technique of base-specific chemical cleavage of Maxam and Gilbert (1980). The nucleotide sequence of araC gene was confirmed by the partial amino acid sequences of araC protein and its methionine peptides. The primary structure of araC polypeptide consists of 291 amino acid residues, giving it a chemical molecular weight of 33 314 daltons. The transcriptional start-point has been deduced from the sequence of araC mRNA synthesized in vitro and in vivo, and it is located 148 bp away from the transcriptional start-point of the araBAD operon. The translational start-point for the araC gene was deduced from the N-terminal amino acid sequence of the protein, and is located 165 bp from the 5'-end of araC mRNA. There is, therefore, a leader sequence of 164 bp preceding the araC gene.

Arabinose↗
PubMed1980

Structure-activity relationships in the development of hypoxic cell radiosensitizers. III. Effects of basic substituents in nitroimidazole sidechains.

The effects of substituting 2-nitroimidazoles with groups carrying basic functions were studied. Prototropic, redox, lipophilicity and protein-binding properties were compared with the efficiency in radiosensitizing hypoxic Chinese hamster V79-379A cells in vitro and the cytotoxicities of the compounds after chronic aerobic exposure. Seventeen compounds were (2-nitro-1-imidazolyl)alkylamines in which the effects of changes in the terminating base and of alkyl chain length were investigated. About an order of magnitude increase in sensitization efficiency could be observed in some compounds without any increase in the aerobic cytotoxicity compared to simple, uncharged 2-nitroimidazoles such as misonidazole. The behaviour of five hydrazones was similar to that of uncharged analogues. The methiodide quaternary salts of two of the (2-nitro-1-imidazolyl)alkylamines showed that quaternization considerably reduced sensitization efficiency. (Nitro-1-imidazolyl)alkylamines appear worthy of further investigation as hypoxic cell radiosensitizers in vivo.

Animals↗
PubMed1980

Toxicity of nitro compounds toward hypoxic mammalian cells in vitro: dependence on reduction potential.

Fifteen nitroaromatic and nitroheterocyclic compounds that can act as "radiosensitizers" were tested for their cytotoxicity toward hypoxic Chinese hamster V79 cells in vitro. The cytotoxicity increased markedly as the electron affinity, measured as a one-electron reduction potential, increased. Non-nitro-containing compounds of similar electron affinities (such as quinones) that also act as radiosensitizers did not exhibit this specific toxicity toward hypoxic cells. The implications of the presence of the nitro group as a prerequisite for the hypoxic cell toxicity were discussed, and the mechanism of the cytotoxicity was compared with that of hypoxic cell radiosensitization.

Animals↗
PubMed1979

Structure-activity relationships in the development of hypoxic cell radiosensitizers. I. Sensitization efficiency.

The efficiency of 35 nitroaromatic and nitroheterocyclic compounds in radiosensitizing hypoxic Chinese Hamster cells in vitro was determined. The concentration C of the compound required to achieve an enhancement ratio of 1.6 was measured, and the redox and partition properties were quantified as the one-electron reduction potential at pH 7, E, and the octanol: water partition coefficient, P, respectively. Most of the compounds studied were 2-nitroimidazoles, but some 4- and 5-nitromidazoles, 5-nitrofurans and nitrobenzenes were investigated for comparison. Together with data for nine nitroimidazoles previously reported, the results were fitted to a structure-activity relationship of the form -log C = b0 + b1E + b2 log P + b3 (log P)2 using multiple linear regression analysis. Statistical tests showed that the coefficients b2 and b3 were not significantly different from zero and the simpler equation, obtained by omitting the terms in log P, explained 85 per cent of the variance in log C. Earlier reports that the radiosensitization efficiency of nitro compounds in vitro largely depends on the reduction potential were confirmed. The conclusive demonstration that P is unimportant in vitro is valuable in interpreting the results of experiments in vivo, where P is expected to have a much greater influence on biological response.

Animals↗
PubMed1979

Structure-activity relationships in the development of hypoxic cell radiosensitizers. II. Cytotoxicity and therapeutic ratio.

This paper describes measurements of the aerobic cytotoxicity of 42 nitroaromatic and nitroheterocyclic compounds towards Chinese Hamster cells in vitro. The results of acute and chronic exposure were quantified, and the concentration C required to achieve a standard response estimated. Fitting the data to an equation of the form - log C = b0 + b1E, where E is the one-electron reduction potential, explained 47 and 71 per cent of the variance in the acute and chronic aerobic cytotoxicity respectively. The addition of further terms to the equation, quantifying partition properties, was not statistically significant. The coefficient b1 was similar for both acute and chronic exposure; the dependence of both cytotoxicity and radiosensitization efficiency on reduction potential was also similar. A therapeutic ratio derived from these in vitro measurements showed no dependence on redox or partition properties. The insensitivity of cytotoxicity and radiosensitization properties to variations in molecular structure, other than those which influence redox behaviour, offers exceptional flexibility in developing compounds of improved therapeutic ratio.

Animals↗
PubMed1978

The rationale for the development of improved hypoxic cell radiosensitizers.

This paper outlines the chemical background to the selection of compounds likely to be clinically superior to the nitroimidazoles currently being evaluated in cancer therapy. The most important chemical properties to consider are electron affinity and lipophilicity, and the quantification and prediction of both these properties are considered. Examples are presented of the use of multiple regression analysis to evaluate the relative contribution of individual properties to the overall biological response.

Nitroimidazoles↗
PubMedDate not supplied

Radiosensitization by non-nitro compounds.

The effects of 23 non-nitro compounds on the radiosensitivity of hypoxic Chinese hamster V79-379A or E. coli AB 1157 cells in vitro are outlined. Imidazole derivatives substituted with several alternative electron-withdrawing groups are described; the dicyanovinyl function conferred considerable radiosensitizing activity. 2,4,5-Tribromoimidazole and 2,4-dinitrophenol may show unusual radiosensitizing activity because of interference with oxidative phosphorylation. Attempts to influence radiosensitivity by compounds potentially capable of depleting intracellular sulphydryls are also described.

Animals↗
PubMedDate not supplied

Bile salt roles in bile-salt-stimulated lipase activity.

The hydrolysis of 4-nitrophenylacetate and phenylsalicylate, catalyzed by human milk lipase in the presence of a range of concentrations of sodium cholate, has been measured at pH 7.3 and 37.5 degrees C, and maximum activity was observed for both substrates at 1 mmole/dm-3 bile salt. Lineweaver-Burk plots for the enzyme-catalyzed hydrolysis of N-methylindoxyl myristate and 4-nitrophenyloctanoate yielded values of Km equal to 34 and 20 mumoles/dm-3, respectively. However, an increase in the concentration of the latter substrate beyond 10 mumoles/dm-3 was not accompanied by a corresponding increase in the rate of hydrolysis. Comparison of these hydrolysis data with literature data for a variety of hydrophobic substrates suggests that there are two roles for the bile salt in the enzyme-catalyzed reaction--the first involving binding onto several sites on the enzyme, and the second solubilization of oil-phase substrates.

Bile Acids and Salts↗