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Literature summary for 6.5.1.2 extracted from

  • Sriskanda, V.; Shuman, S.
    A second NAD+-dependent DNA ligase (LigB) in Escherichia coli (2001), Nucleic Acids Res., 29, 4930-4934.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 6.5.1.2

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21 (DE3) Escherichia coli

Protein Variants

Protein Variants Comment Organism
K126A inactive Escherichia coli

Inhibitors

Inhibitors Comment Organism Structure
NAD+ inhibitory above 0.1 mM, activity is abolished at 0.5 mM Escherichia coli

Metals/Ions

Metals/Ions Comment Organism Structure
Co2+ 25% of the activity with Mg2+ Escherichia coli
Mg2+ 1-10 mM, required Escherichia coli
Mn2+ 1-10 mM, required Escherichia coli
Zn2+ no effect Escherichia coli

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
NAD+ + (deoxyribonucleotide)n + (deoxyribonucleotide)m Escherichia coli DNA-repair, phosphodiester bond formation between adjacent 5'-phosphate and 3'-hydroxyl groups in double-stranded DNA AMP + nicotinamide nucleotide + (deoxyribonucleotide)n+m
-
 ?

Organism

Organism UniProt Comment Textmining
Escherichia coli
-
 second NAD+-dependent ligase LigB, high sequence homology to LigA, lacks zinc binding motif that is found in LigA
-

Purification (Commentary)

Purification (Comment) Organism
recombinant protein using His-tag Escherichia coli

Reaction

Reaction Comment Organism Reaction ID
ATP + (deoxyribonucleotide)n-3'-hydroxyl + 5'-phospho-(deoxyribonucleotide)m = (deoxyribonucleotide)n+m + AMP + beta-nicotinamide D-nucleotide 3 steps of reaction: 1. adenylation of the ligase in the presence of NAD+, 2. transferring the adenylate moiety to the 5'-phosphate of the nicked DNA substrate, 3. sealing the nick through the formation of a phosphodiester bond Escherichia coli
a

Storage Stability

Storage Stability Organism
-80°C Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + nicked DNA
-
 Escherichia coli AMP + diphosphate + ?
-
 ?
NAD+ + (deoxyribonucleotide)n + (deoxyribonucleotide)m
-
 Escherichia coli AMP + nicotinamide nucleotide + (deoxyribonucleotide)n+m
-
 ?
NAD+ + (deoxyribonucleotide)n + (deoxyribonucleotide)m DNA-repair, phosphodiester bond formation between adjacent 5'-phosphate and 3'-hydroxyl groups in double-stranded DNA Escherichia coli AMP + nicotinamide nucleotide + (deoxyribonucleotide)n+m
-
 ?

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0008
-
 nicked DNA
-
 Escherichia coli

pH Range

pH Minimum pH Maximum Comment Organism
6.5 9
-
 Escherichia coli

Cofactor

Cofactor Comment Organism Structure
NAD+ required, highest activity 0.007-0.06 mM Escherichia coli