scholarly journals The schiff base complex of yeast 5-aminolaevulinic acid dehydratase with laevulinic acid

1999 ◽  
Vol 8 (6) ◽  
pp. 1250-1256 ◽  
Author(s):  
Peter T. Erskine ◽  
Richard Newbold ◽  
Jenny Roper ◽  
Alun Coker ◽  
Martin J. Warren ◽  
...  
Keyword(s):  
Schiff Base ◽  
Schiff Base Complex ◽  
Base Complex ◽  
Aminolaevulinic Acid ◽  
Acid Dehydratase ◽  
Aminolaevulinic Acid Dehydratase

scholarly journals Characterization of the two 5-aminolaevulinic acid binding sites, the A- and P-sites, of 5-aminolaevulinic acid dehydratase from Escherichia coli

1995 ◽  
Vol 305 (1) ◽  
pp. 151-158 ◽  
Author(s):  
P Spencer ◽  
P M Jordan
Keyword(s):  
Schiff Base ◽  
Binding Sites ◽  
Metal Ion ◽  
Substrate Binding ◽  
Aminolaevulinic Acid ◽  
Acid Dehydratase ◽  
Carboxylate Groups ◽  
Catalytic Metal ◽  
A Site ◽  
Aminolaevulinic Acid Dehydratase

Experiments are described in which the individual properties of the two 5-aminolaevulinic acid (ALA) binding sites, the A-site and the P-site, of 5-aminolaevulinic acid dehydratase (ALAD) have been investigated. The ALA binding affinity at the A-site is greatly enhanced (at least 10-fold) on the binding of the catalytic metal ion (bound at the alpha-site). The nature of the catalytic metal ion, Mg2+ or Zn2+, also gave major variations in the substrate Km, P-site affinity for ALA, the effect of potassium and phosphate ions and the pH-dependence of substrate binding. Modification of the P-site by reaction of the enzyme-substrate Schiff base with NaBH4 and analysis of the reduced adduct by electro-spray mass spectrometry indicated a maximum of 1 mol of substrate incorporated/mol of subunit, correlating with a linear loss of enzyme activity. The reduced Schiff-base adduct was used to investigate substrate binding at the A-site by using rate-of-dialysis analysis. The affinity for ALA at the A-site of Mg alpha Zn beta ALAD was found to determine the Km for the reaction and was pH-dependent, with its affinity increasing from 1 mM at pH 6 to 70 microM at pH 8.5. The affinity of ALA at the P-site of Zn alpha An beta ALAD is proposed to limit the Km at pH values above 7, since the measured Kd for ALA at the A-site in 45 microM Tris, pH 8, was well below the observed Km (600 microM) under the same conditions. The amino group of the ALA molecule bound at the P-site was identified as a critical binding component for the A-site, explaining why ALA binding to ALAD is ordered, with the P-site ALA binding first. Structural requirements for ALA binding at the A- and P-sites have been identified: the P-site requires the carbonyl and carboxylate groups, whereas the A-site requires the amino, carbonyl and carboxylate groups of the substrate.

5-Aminolaevulinic acid dehydratase: structure, function, and mechanism

1976 ◽  
Vol 273 (924) ◽  
pp. 109-115 ◽  
Keyword(s):  
Schiff Base ◽  
Molecular Mass ◽  
Active Sites ◽  
Bovine Liver ◽  
Sh Groups ◽  
Aminolaevulinic Acid ◽  
Acid Dehydratase ◽  
Arginine Residues ◽  
Assay Conditions ◽  
Aminolaevulinic Acid Dehydratase

S-Aminolaevulinic acid dehydratase catalyses the synthesis of porphobilinogen. The enzyme has a molecular mass of 285000 and is composed of eight similar subunits of molecular mass 35000. The N-terminal amino acid is acylated, and the number of peptides found on tryptic digestion equals the number of lysine and arginine residues per mass of 35000. The eight subunits are apparently arranged at the corners of a cube and therefore have dihedral (D 4 ) symmetry. The bovine liver enzyme which has been crystallized contains 4-6 atoms of zinc per mole of enzyme. The apo-enzyme obtained on prolonged hydrolysis can be reactivated by the addition of zinc or cadmium ions. The dialysed enzyme must be first treated with dithiothreitol. There are two very active SH groups in a total of 6-7-SH groups per subunit. The substrate forms a Schiff base with the e-amino group of a lysine residue. Reduction of the Schiff base with NaBH 4 should reveal the number of active sites per mole of enzyme. It appears that only four of the eight subunits form a Schiff base with the substrate indicating that the enzyme exhibits the phenomenon of either half-site reactivity or negative cooperativity. The enzyme appears to have a strong subunit-subunit interaction for an immobilized preparation remained stable for at least a month. An immobilized enzyme preparation was treated in a manner so that it dissociated into tetramers. Both the eluate and the protein still attached to the Sepharose on a column were enzymically active. The bound enzyme could not reassociate under assay conditions but still contained about 50 % of the original enzyme activity. It would seem that the enzyme is active when composed with less than eight subunits.

The DNA- and protein-binding properties and cytotoxicity of a new copper(II) hydrazone Schiff base complex

2021 ◽  
pp. 1-23
Author(s):  
Niladri Biswas ◽  
Sandeepta Saha ◽  
Barun Kumar Biswas ◽  
Manas Chowdhury ◽  
Ashikur Rahaman ◽  
...  
Keyword(s):  
Schiff Base ◽  
Protein Binding ◽  
Schiff Base Complex ◽  
Binding Properties ◽  
Base Complex
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