Magnesium Activation of Ribonuclease H. Evidence for One Catalytic Metal Ion

1994 ◽  
Vol 33 (25) ◽  
pp. 5805-5808 ◽  
Author(s):  
C. B. Black ◽  
J. A. Cowan
Keyword(s):  
Metal Ion ◽  
Ribonuclease H ◽  
Catalytic Metal

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.

scholarly journals Structural intermediates of a DNA–ligase complex illuminate the role of the catalytic metal ion and mechanism of phosphodiester bond formation

2019 ◽  
Vol 47 (14) ◽  
pp. 7147-7162 ◽  
Author(s):  
Adele Williamson ◽  
Hanna-Kirsti S Leiros
Keyword(s):  
Active Site ◽  
Metal Ion ◽  
Bond Formation ◽  
Strand Break ◽  
Nucleophilic Attack ◽  
Dna Ligase ◽  
Phosphodiester Bond ◽  
Dna Ligases ◽  
Catalytic Metal ◽  
Ternary Structure

Abstract DNA ligases join adjacent 5′ phosphate (5′P) and 3′ hydroxyl (3′OH) termini of double-stranded DNA via a three-step mechanism requiring a nucleotide cofactor and divalent metal ion. Although considerable structural detail is available for the first two steps, less is known about step 3 where the DNA-backbone is joined or about the cation role at this step. We have captured high-resolution structures of an adenosine triphosphate (ATP)-dependent DNA ligase from Prochlorococcus marinus including a Mn-bound pre-ternary ligase–DNA complex poised for phosphodiester bond formation, and a post-ternary intermediate retaining product DNA and partially occupied AMP in the active site. The pre-ternary structure unambiguously identifies the binding site of the catalytic metal ion and confirms both its role in activating the 3′OH terminus for nucleophilic attack on the 5′P group and stabilizing the pentavalent transition state. The post-ternary structure indicates that DNA distortion and most enzyme-AMP contacts remain after phosphodiester bond formation, implying loss of covalent linkage to the DNA drives release of AMP, rather than active site rearrangement. Additionally, comparisons of this cyanobacterial DNA ligase with homologs from bacteria and bacteriophage pose interesting questions about the structural origin of double-strand break joining activity and the evolution of these ATP-dependent DNA ligase enzymes.

Catalytic Metal Ion Binding in Enolase: The Crystal Structure of an Enolase-Mn2+-Phosphonoacetohydroxamate Complex at 2.4-.ANG. Resolution,

Biochemistry ◽  
1994 ◽  
Vol 33 (20) ◽  
pp. 6295-6300 ◽  
Author(s):  
Erli Zhang ◽  
Marcos Hatada ◽  
John M. Brewer ◽  
Lukasz Lebioda
Keyword(s):  
Crystal Structure ◽  
Metal Ion ◽  
Ion Binding ◽  
Metal Ion Binding ◽  
Catalytic Metal

Diameter control of carbon nanotubes by changing the concentration of catalytic metal ion solutions

Carbon ◽  
2005 ◽  
Vol 43 (7) ◽  
pp. 1453-1459 ◽  
Author(s):  
Lily Kim ◽  
Eun-mi Lee ◽  
Shin-Je Cho ◽  
Jung Sang Suh
Keyword(s):  
Carbon Nanotubes ◽  
Metal Ion ◽  
Catalytic Metal ◽  
Diameter Control

Role of the Divalent Metal Ion in Sugar Binding, Ring Opening, and Isomerization by D-Xylose Isomerase: Replacement of a Catalytic Metal by an Amino Acid

Biochemistry ◽  
1994 ◽  
Vol 33 (6) ◽  
pp. 1488-1494 ◽  
Author(s):  
Karen N. Allen ◽  
Arnon Lavie ◽  
Arthur Glasfeld ◽  
Timothy N. Tanada ◽  
Daniel P. Gerrity ◽  
...  
Keyword(s):  
Amino Acid ◽  
Ring Opening ◽  
Metal Ion ◽  
Xylose Isomerase ◽  
Divalent Metal ◽  
Sugar Binding ◽  
Divalent Metal Ion ◽  
Catalytic Metal
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