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Literature summary extracted from

  • Prigozhin, D.M.; Mavrici, D.; Huizar, J.P.; Vansell, H.J.; Alber, T.
    Structural and biochemical analyses of Mycobacterium tuberculosis N-acetylmuramyl-L-alanine amidase Rv3717 point to a role in peptidoglycan fragment recycling (2013), J. Biol. Chem., 288, 31549-31555.
    View publication on PubMedView publication on EuropePMC

Application

EC Number Application Comment Organism
3.5.1.28 drug development the enzyme can aid in the development of cell wall targeting therapeutics against Mycobacterium tuberculosis Mycobacterium tuberculosis

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.5.1.28 gene Rv3717, expression of recombinant GST-tagged enzyme in Escherichia coli strain BL-21 Codon Plus Mycobacterium tuberculosis

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
3.5.1.28 purified enzyme Rv3717 free, or in complex with L-Ala-iso-D-Gln product or Zn2+, hanging drop vapor diffusion method, from 200 mM NaCl, 100 mM Tris, pH 8.5, 25% PEG 3350, X-ray diffraction structure determination and analysis at 2.1-2.67 A resolution, molecular replacement method using Bartonella henselae AmiB as the search model, PDB code 3NE8 Mycobacterium tuberculosis

Protein Variants

EC Number Protein Variants Comment Organism
3.5.1.28 E200X inactive mutant Mycobacterium tuberculosis

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.5.1.28 Zn2+ Zn2 is required for hydrolysis of muramyl dipeptide, Zn2 binding rearranges active site residues. The catalytic residue Glu200 does not participate in binding to Zn2+ ions Mycobacterium tuberculosis

Organism

EC Number Organism UniProt Comment Textmining
3.5.1.28 Mycobacterium tuberculosis I6Y4D2
-
-
3.5.1.28 Mycobacterium tuberculosis H37Rv I6Y4D2
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.5.1.28 recombinant GST-tagged enzyme from Escherichia coli strain BL-21 Codon Plus by glutathione affinity chromatography and tag cleavage, followed by a combined glutathione affinity/anion exchange chromatography step, and a gel filtration step Mycobacterium tuberculosis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.5.1.28 additional information no activity on polymerized peptidoglycan Mycobacterium tuberculosis ?
-
 ?
3.5.1.28 additional information no activity on polymerized peptidoglycan Mycobacterium tuberculosis H37Rv ?
-
 ?
3.5.1.28 MurNAc-L-Ala-D-isoGln + H2O
-
 Mycobacterium tuberculosis N-acetylmuramic acid + L-alanyl-iso-D-glutamine
-
 ?
3.5.1.28 MurNAc-L-Ala-D-isoGln + H2O
-
 Mycobacterium tuberculosis H37Rv N-acetylmuramic acid + L-alanyl-iso-D-glutamine
-
 ?

Synonyms

EC Number Synonyms Comment Organism
3.5.1.28 Mtb peptidoglycan amidase
-
 Mycobacterium tuberculosis
3.5.1.28 Rv3717
-
 Mycobacterium tuberculosis

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.5.1.28 22
-
 assay at room temperature Mycobacterium tuberculosis

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.5.1.28 6.5
-
 assay at Mycobacterium tuberculosis

General Information

EC Number General Information Comment Organism
3.5.1.28 evolution the enzyme belongs to the Pfam family Amidase_3, the mode of substrate binding is likely shared by the Amidase_3 family proteins Mycobacterium tuberculosis
3.5.1.28 additional information the enzyme contains an extra disulfide-bonded beta-hairpin adjacent to the active site compared to other peptidoglycan amidases. Zn2 binding rearranges active site residues and disulfide formation promotes folding of the beta-hairpin. Although Zn2 is required for hydrolysis of muramyl dipeptide, disulfide oxidation is not required for activity on this substrate. The product L-alanine-iso-D-glutamine binds at the head of a closed tunnel. The orientation of the product in the active site suggests a role for a conserved Glu200 in catalysis Mycobacterium tuberculosis
3.5.1.28 physiological function potential role for enzyme Rv3717 in peptidoglycan fragment recycling Mycobacterium tuberculosis