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Literature summary extracted from

  • Paul, A.; Kumar, P.; Surolia, A.; Vijayan, M.
    Biochemical and structural studies of mutants indicate concerted movement of the dimer interface and ligand-binding region of Mycobacterium tuberculosis pantothenate kinase (2017), Acta Crystallogr. Sect. F, 73, 635-643 .
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.1.33 gene coaX, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 Mycobacterium tuberculosis

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
2.7.1.33 purified recombinant His-tagged wild-type and mutant enzymes in apoform, microbatch-under-oil method, mixing of 0.002 ml of 6.5 mg/ml protein in 100 mM Tris-HCl, pH 7.8, 600 mM NaCl, 5% glycerol, and 100 mM imidazole, with 0.002 ml of precipitant solution that contains for mutant F247A: 0.2 M (NH4)2SO4, 0.1 M bis-Tris, pH 5.5, and 25% w/v PEG 3350, for mutant F254A: 10-15% w/v PEG 400, 1.25 M (NH4)2SO4, and 100 mM Na HEPES, pH 7.5, and for the double mutant F247A/F254A: 2% w/v PEG 400, 2.0 M (NH4)2SO4, and 100 mM Na HEPES, pH 7.5, at 22°C, X-ray diffraction structure determination and analysis at 1.80-3.20 A resolution, modeling Mycobacterium tuberculosis

Protein Variants

EC Number Protein Variants Comment Organism
2.7.1.33 F247A site-directed mutagenesis, determination of the crystal structure and comparion with the wild-type enzyme structure and the structure of Escherichia coli enzyme EcPanK Mycobacterium tuberculosis
2.7.1.33 F254A site-directed mutagenesis, determination of the crystal structure and comparion with the wild-type enzyme structure and the structure of Escherichia coli enzyme EcPanK Mycobacterium tuberculosis
2.7.1.33 F254A/F247A site-directed mutagenesis, determination of the crystal structure and comparion with the wild-type enzyme structure and the structure of Escherichia coli enzyme EcPanK Mycobacterium tuberculosis
2.7.1.33 additional information generation of two point mutants and the corresponding double mutant of Mycobacterium tuberculosis pantothenate kinase to weaken the affinity of the enzyme for the feedback inhibitor CoA. The mutants exhibit reduced activity, which can be explained in terms of their structures, structure-function analysis, overview. The crystals of the mutants are not isomorphous to any of the previously analysed crystals of the wild-type enzyme or its complexes. Although the mutants involve changes in the CoA-binding region, the dimer interface and the ligand-binding region move in a concerted manner, an observation which might be important in enzyme action. The mycobacterial enzyme and its homologous Escherichia coli enzyme exhibit structural differences in their nucleotide complexes in the dimer interface and the ligand-binding region, but in three of the four crystallographically independent mutant molecules the structure is similar to that in the Escherichia coli enzyme Mycobacterium tuberculosis

General Stability

EC Number General Stability Organism
2.7.1.33 MtPanK is a robust enzyme Mycobacterium tuberculosis

Inhibitors

EC Number Inhibitors Comment Organism Structure
2.7.1.33 CoA feedback inhibition, Tm values of wild-type and its mutants and dissociation constants of CoA, overview Mycobacterium tuberculosis

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.1.33 Mg2+ required Mycobacterium tuberculosis

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.7.1.33 ATP + (R)-pantothenate Mycobacterium tuberculosis
-
 ADP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 ATP + (R)-pantothenate Mycobacterium tuberculosis H37Rv
-
 ADP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 ATP + (R)-pantothenate Mycobacterium tuberculosis ATCC 25618
-
 ADP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 GTP + (R)-pantothenate Mycobacterium tuberculosis
-
 GDP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 GTP + (R)-pantothenate Mycobacterium tuberculosis H37Rv
-
 GDP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 GTP + (R)-pantothenate Mycobacterium tuberculosis ATCC 25618
-
 GDP + (R)-4'-phosphopantothenate
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.7.1.33 Mycobacterium tuberculosis P9WPA1
-
-
2.7.1.33 Mycobacterium tuberculosis ATCC 25618 P9WPA1
-
-
2.7.1.33 Mycobacterium tuberculosis H37Rv P9WPA1
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.1.33 recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 by nickel affinity chromatography and dialysis Mycobacterium tuberculosis

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.1.33 ATP + (R)-pantothenate
-
 Mycobacterium tuberculosis ADP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 ATP + (R)-pantothenate
-
 Mycobacterium tuberculosis H37Rv ADP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 ATP + (R)-pantothenate
-
 Mycobacterium tuberculosis ATCC 25618 ADP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 GTP + (R)-pantothenate
-
 Mycobacterium tuberculosis GDP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 GTP + (R)-pantothenate
-
 Mycobacterium tuberculosis H37Rv GDP + (R)-4'-phosphopantothenate
-
 ?
2.7.1.33 GTP + (R)-pantothenate
-
 Mycobacterium tuberculosis ATCC 25618 GDP + (R)-4'-phosphopantothenate
-
 ?

Subunits

EC Number Subunits Comment Organism
2.7.1.33 dimer
-
 Mycobacterium tuberculosis

Synonyms

EC Number Synonyms Comment Organism
2.7.1.33 CoaX
-
 Mycobacterium tuberculosis
2.7.1.33 MtPanK
-
 Mycobacterium tuberculosis
2.7.1.33 PanK
-
 Mycobacterium tuberculosis

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
2.7.1.33 35.7
-
 Tm of enzyme mutant F254A without CoA Mycobacterium tuberculosis
2.7.1.33 36.7
-
 Tm of enzyme mutant F247A/F254A without CoA Mycobacterium tuberculosis
2.7.1.33 37
-
 Tm of enzyme mutant F247A without CoA Mycobacterium tuberculosis
2.7.1.33 40.1
-
 Tm of enzyme mutant F247A/F254A with 0.2 mM CoA Mycobacterium tuberculosis
2.7.1.33 40.1
-
 Tm of enzyme mutant F254A with 0.2 mM CoA Mycobacterium tuberculosis
2.7.1.33 42.4
-
 Tm of enzyme mutant F247A with 0.2 mM CoA Mycobacterium tuberculosis
2.7.1.33 43.8
-
 Tm of wild-type enzyme without CoA Mycobacterium tuberculosis
2.7.1.33 50.1
-
 Tm of wild-type enzyme with 0.2 mM CoA Mycobacterium tuberculosis
2.7.1.33 80
-
 purified recombinant enzyme, 10 min, pH 8.0, inactivation Mycobacterium tuberculosis

General Information

EC Number General Information Comment Organism
2.7.1.33 evolution comparison of MtPanK with the Escherichia coli enzyme EcPanK, overview. Despite the high sequence identity (52%) between EcPanK and MtPanK, the two enzymes differ in many respects, crystal structure comparison. While EcPanK is specific for ATP, MtPanK exhibits dual specificity and can make use of ATP as well as GTP for phosphorylating pantothenic acid. CoA binds nearly 40% more tightly to MtPanK than to EcPanK Mycobacterium tuberculosis
2.7.1.33 metabolism the biosynthesis of CoA consists of five enzymatically catalysed steps, the first of which involves the conversion of vitamin B5 (pantothenate) to 4'-phosphopantothenate by ATP-mediated phosphorylation carried out by pantothenate kinase Mycobacterium tuberculosis