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Biomedical subjects

M Farstad

Publications and source records attributed to M Farstad.

At least 73 records · Page 4Linked to original sources
PubMed1981

Isolation of palmitoyl-CoA hydrolases from human blood platelets.

The palmitoyl-CoA hydrolase activity, which in human blood platelets is mainly localized in the cytosol fraction [Berge, Vollset & Farstad (1980) Scand. J. Clin. Lab. Invest. 40, 271--279], was found to be extremely labile. Inclusion of glycerol or palmitoyl-CoA stabilized the activity during preparation. Gel-filtration studies revealed multiple forms of the enzyme with molecular weights corresponding to about 70 000, 40 000 and 24 000. The relative recovery of the mol.wt.-70 000 form was increased by the presence of 20% (v/v) glycerol or 10 microM-palmitoyl-CoA. The three enzyme forms are probably unrelated, since they were not interconvertible. The three different species of palmitoyl-CoA hydrolase were purified by DEAE-cellulose and hydroxyapatite chromatography, isoelectric focusing and high-pressure liquid chromatography (h.p.l.c.) to apparent homogeneity. The three enzymes had isoelectric points (pI) of 7.0, 6.1 and 4.9. The corresponding molecular weights were 27 000--33 000, 66 000--72 000 and 45 000--49 000, calculated from h.p.l.c. and Ultrogel AcA-44 chromatography. The apparently purified enzymes were unstable, as most of the activity was lost during purification. The enzyme with an apparent molecular weight of 45 000--49 000 was split into fractions with molecular weights of less than 10 000 by re-chromatography on h.p.l.c. concomitantly with a loss of activity. The stimulation of the activity by the presence of serum albumin seems to depend on the availability of palmitoyl-CoA, as has been reported for other palmitoyl-CoA hydrolases. [Berge & Farstad (1979) Eur. J. Biochem. 96, 393--401].

Blood Platelets↗
PubMed1981

Variations in the activity of microsomal palmitoyl-CoA hydrolase in mixed micelle solutions of palmitoyl-CoA and non-ionic detergents of the triton X series.

The kinetics of palmitoyl-CoA hydrolase were influenced by both the availability of the substrate and formation of micelles. At palmitoyl-CoA concentrations below the critical micelle concentration, addition of non-ionic detergent increased the activity until the critical micelle concentration of the mixed micelles was reached. At palmitoyl-CoA concentrations above the critical micelle concentration, inhibitor of the activity was observed, but addition of detergents of the Triton X series reversed the inhibition. Maximum palmitoyl-CoA hydrolase activity was found when the ratios (w/v) of palmitoyl-CoA: Triton X-100 and palmitoyl-CoA: Triton X-405 were approximately 0.35 and 0.05, respectively. At these above the mixed critical micelle concentration. The results indicate that monomer palmitoyl-CoA is the substrate and that monomer forms of the non-ionic detergents of the Triton X series activate the enzyme. Isolated microsomal lipids activated the microsomal palmitoyl-CoA hydrolase, suggesting that a hydrophobic environment is advantageous for interaction between enzyme and substrate in vivo. The maximum activity in the presence of mixed micelles is discussed in relation to a model where mixed micelles are regarded as artificial membranes to which the enzyme may adhere in an equilibrium with the monomer substrate and detergent in the monomer form. It is suggested that intracellular membranes may resemble mixed micelles in equilibrium with detergent-active substrates such as palmitoyl-CoA.

Animals↗
PubMed1980

Direct measurement of free coenzyme A in biological extracts by reversed-phase high-performance liquid chromatography.

A high-performance liquid chromatographic system was developed with baseline separation of coenzyme A (CoASH) from dephospho-coenzyme A and acetyl-coenzyme A using isocratic elution. The chromatographic separation was achieved in a reversed-phase system with a high concentration (220 mM) of potassium phosphate buffer at pH 4.0 and appropriate amounts of methanol (ca. 12%). The eluate was monitored with a UV detector at 254 nm with the limit of detection at ca. 5 pmoles. The system could be used without modification for the estimation of the content of CoASH in biological extracts, e.g. mitochondria.

Animals↗
PubMed1980

Proteinuria after selective nephroangiography in man. Comparison of three contrast media.

The proteinuria following aortography and selective nephroangiography in 60 patients using Isopaque Cerebral, Urografin 60% and Amipaque was investigated. The results indicate an increase in glomerular permeability and possibly an overload of tubular reabsorptive capacity after angiography. The changes are transient. No permanent injury to the kidney appears to occur.

Albuminuria↗
PubMed1980

Intracellular localization of palmitoyl-CoA hydrolase and palmitoyl-CoA synthetase in human blood platelets and liver.

The intracellular localization studies of human blood platelets by sucrose gradient and differential centrifugation, showed that palmitoyl-CoA hydrolase was mainly localized in the cytosol fraction. However, a localization also in the mitochondrial fraction seems possible as disruption of platelets by the French press and nitrogen decompression techniques resulted in mitochondrial damage. In human liver the palmitoyl-CoA hydrolase was localized in the mitochondrial and microsomal fractions. Palmitoyl-CoA synthetase was localized in the mitochondrial and microsomal fractions in both platelets and liver. The possible physiological implications of these differences, and the finding of a very high ratio of palmitoyl-CoA hydrolase/palmitoyl-CoA synthetase in platelets compared with liver, are discussed.

Blood Platelets↗
PubMed1979

Intracellular localization of long-chain acyl-coenzyme A hydrolase and acyl-L-carnitine hydrolase in brown adipose tissue from guinea pigs.

The activities of long-chain acyl-CoA hydrolase (palmitoyl-CoA hydrolase, EC 3.1.2.2) and long-chain acyl-L-carnitine hydrolase, EC 3.1.1.28) in brown adipose tissue from cold-exposed and control guinea pigs were studied. Mitochondria from cold-exposed animals hydrolysed 21 nmol of palmitoyl-CoA/min per mg of protein and 1.3 nmol of palmitoyl-L-carnitine/min per mg of protein, and the specific activities were respectively 2 and 5 times as high in cold-exposed as in control animals. The subcellular-localization studies showed that both the long-chain acyl-CoA hydrolase and long-chain acyl-L-carnitine hydrolase were localized in the mitochondria. A location also in the soluble fraction cannot be excluded. The long-chain acyl-CoA hydrolase activity was doubled when the mitochondria were disrupted; this indicates that the enzyme is localized in the matrix compartment.

Adipose Tissue, Brown↗
PubMed1979

Dual localization of long-chain acyl-CoA hydrolase in rat liver: one in the microsomes and one in the mitochondrial matrix.

Subcellular fractionation studies of rat liver localized the activity of palmitoyl-L-carnitine hydrolase to the microsomal fraction whereas palmitoyl-CoA hydrolase activity was found both in the microsomal fraction and in mitochrondria. An unusual biphasic sataration curve for palmitoyl-CoA was observed when intact mitochondrial hydrolase activity. Disruption of the mitochondrial structure doubled the palmitoyl-CoA hydrolysis. Discontinuous sucrose gradient centrifugation and digitonin fractionation of rat liver mitochondria demonstrated that a palmitoyl-CoA hydrolase was associated with the matrix fraction. Pure matrix and microsomal fractions showed that the two hydrolase activities were differently affected by the presence of divalent cations. Both the specific activity and the saturation concentration of palmitoyl-CoA were higher for the microsomal enzyme than for the matrix-associated enzyme.

Animals↗
PubMed1979

A study of assay conditions for palmitoyl-CoA synthetase and carnitine palmitoyltransferase in homogenates of human blood platelets.

The assay conditions for palmitoyl-CoA synthetase (P-CoA S) and carnitine palmitoyltransferase (CPT) in homogenates of human blood platelets have been studied. The assay based on trapping of palmitoyl-CoA by carnitine in the presence of exogenous CPT gave higher activity of P-CoA S than the assay based on direct isolation of the palmitoyl-CoA formed. The activity of CPT was higher on exogenous palmitoyl-CoA than on endogenous palmitoyl-CoA formed from palmitic acid and CoA in the presence of endogenous P-CoA S. The activity of CPT was strongly dependent on the incubation time and the amount of platelets used. The initial activity of this enzyme in human blood platelets was higher than previously assumed.

Acyltransferases↗