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Biomedical subjects

F Lynen

Publications and source records attributed to F Lynen.

At least 37 records · Page 2Linked to original sources
PubMed1977

Reaction of yeast fatty acid synthetase with iodoacetamide. 3. Malonyl-coenzyme A decarboxylase as product of the reaction of fatty acid synthetase with iodoacetamide.

Yeast fatty acid synthetase possesses very low malonyl-CoA decarboxylase activity. Treatment with iodoacetamide, while abolishing synthetase activity, induces a strong malonyl decarboxylase activity which, in turn, can be inhibited by N-ethylmaleimide. Kinetic analysis shows that the emergence of the decarboxylase activity is synchronized to the disappearance of the fatty-acid-synthesizing activity and thus, is due to carboxamidomethylation of the peripheral SH-groups of the multienzyme complex. Strong decarboxylase activity was also found after treatment of the synthetase with methylmalonyl-CoA. A hypothetical scheme is proposed which explains the origination of the decarboxylase activity as a consequence of conformational changes of the condensing enzyme component which happen when the peripheral SH-group is acylated or alkylated.

Binding Sites↗
PubMed1976

6-Methylsalicylic acid synthetase from Penicillium patulum. Some catalytic properties of the enzyme and its relation to fatty acid synthetase.

1. The specificity of 6-methylsalicylic acid synthetase with respect to the priming substrate was studied. If acetyl-CoA was replaced by propionyl-CoA 6-ethylsalicylic acid was synthesized. The rate of this synthesis was about 13% that of 6-methylsalicylic acid synthesis in presence of acetyl-CoA. 2. 6-Methylsalicylic acid synthetase contains an acetyl transferase activity as demonstrated by the transfer of acetyl residues from acetyl-CoA to pantetheine. This transferase also catalyses propionyl transfer but only with an about 13 times slower rate. 3. Analogous to fatty acid synthetase, treatment with iodoacetamide converts 6-methylsalicylic acid synthetase into a malonyl-CoA decarboxylase. Under certain conditons the iodoacetamide-treated enzyme could catalyse 6-methylsalicylic acid formation from malonyl CoA and NADPH without the external addition of acetyl CoA.

Acetyl Coenzyme A↗
PubMed1975

Investigations of the structure of 3-methylcrotonyl-CoA carboxylase from Achromobacter.

It was shown by gel electrophoresis in sodium dodecylsulphate solution that 3-methylcrotonyl-CoA carboxylase from Achromobacter IVS is composed of two different subunits with molecular weights of about 78000 and 96000, respectively. The biotin is bound to the heavier subunit. It was previously found that 3-methylcrotonyl-CoA carboxylase contains four biotin molecules per complex. A complex composed of four of each subunit would thus have a molecular weight of about 700000. This is compatible with the molecular weight of 760000 determined earlier by analytical ultracentrifugation. Both subunits were isolated preparatively. As the subunits, unlike the complex, are very sensitive to oxygen, special precautions had to be taken during isolation. The biotin-containing subunit was isolated by chromatography on DEAE-cellulose in 5 M urea. It no longer catalyzed the overall reaction, yet could still carboxylate free biotin. The biotin-free subunit was separated after dissociation of the enzyme by three-days' dialysis at pH 9.8 under nitrogen. On chromatography over a Sepharose-bound avidin column, the biotin-subunit was fixed and the biotin-free subunit was eluted unretarded. The latter subunit showed no enzymic activity. After the addition of the biotin-containing subunit, overall activity was regenerated. The speed of reassociation is very much enhanced by 3-methylcrotonyl-CoA. It was shown by reassociation experiments under different conditions that probably an initial complex, AxBy is formed, possessing a binding site for 3-methylcrotonyl-CoA. Upon the binding of this substrate the conformation may be changed to a form favourable for reconstitution. Finally, the structures of biotin enzymes from different sources are compared. In the course of evolution there is a tendency toward integration of the different constituent proteins into only one polypeptide chain.

Alcaligenes↗
PubMed1975

Patulin biosynthesis: the metabolism of m-hydroxybenzyl alcohol and m-hydroxybenzaldehyde by particulate preparations from Penicillium patulum.

The ring hydroxylation of m-hydroxybenzyl alcohol to gentisyl alcohol by a particulate preparation from Penicillium patulum has been characterised. The activity was shown to be closely associated with, but not necessarily identical to, m-cresol 2-hydroxylase activity of the 105 000 X g microsomal fraction. As with both the m-cresol hydroxylases of this system, m-hydroxybenzyl alcohol hydroxylase required oxygen and NADPH for activity. A Km value for m-hydroxybenzyl alcohol of 15 muM was measured. Inhibition of the hydroxylase activity and its reversal by light, as well as the action of cytochrome c, KCN and other effectors suggested a mixed-function oxidase reaction of the cytochrome P-450, NADPH-cytochrome reductase type. m-Hydroxybenzaldehyde was not ring hydroxylated by any preparation from P. patulum. Apart from the previously described conversion to m-hydroxybenzyl alcohol by a predominantly soluble dehydrogenase, m-hydroxybenzaldehyde was metabolized to m-hydroxybenzoic acid by a particulate fraction. This activity required NADPH. It was concluded that the main biosynthetic pathway to patulin must be through m-hydroxybenzyl alcohol, gentisyl alcohol and gentisaldehyde.

Benzaldehydes↗
PubMed1975

[Mechanism for the condensation reaction of fatty-acid biosynthesis (author's transl)].

The spontaneous hydrogen-deuterium exchange of the methylene group of malonyl-thioesters was investigated by nuclear-magnetic-resonance (NMR) spectroscopy using the model compound S-malonyl-N-acetylcysteamine. The half life of the methylene proteins is 12 to 16 min in 0.1 M K-phosphate buffer at pH 6.5 to 7.0 at 25 degrees C, the conditions of maximal activity of fatty acid synthetase from yeast. Proton catalysis was used for the quick preparation of deuterium- and tritium-labeled malonylthioesters. Compared with malonyl-CoA, dideutero-malonyl-CoA had no primary isotope effect on the reaction velocity of the yeast enzyme catalysed fatty acid synthesis, in which the rate limiting step is the condensation reaction. Although deuterium oxide had a solvent isotope effect, there was no difference in reaction velocities between malonyl CoA and dideuteromalonyl CoA in deuterium oxide. The condensation reaction was investiaged separately from the overall fatty acid synthesis using beta-ketoacyl-acyl-carrier-protein (ACP) synthetase (condensing enzyme) of Escherichia coli. The condensation reaction with deuteromalonyl-ACP had no kinetic isotope effect, in agreement with the observations on the overall reaction. However, in this case no solvent isotope effect was observed with 2H2O. When the condensation reaction was carried out in the presence of tritiated water, there was no incorporation of label into the reaction product acetoacetyl-thioester, excluding proton exchange with the solvent. The results exclude a mechanism for the condensation reaction involving a malonyl carbanion and its acylation as intermediates in the sense of an organic-chemical malonic ester synthesis, and they indicate that the condensation reaction follows a concerted mechanism: The formation of the new carbon-carbon bond is coupled with the cleavage of the carboxyl bond of the malonyl group.

Binding Sites↗