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Shikimate Kinase (shikimate + kinase)

Distribution by Scientific Domains
Chemistry100%

Selected Abstracts

The refolding of type II shikimate kinase from Erwinia chrysanthemi after denaturation in urea

FEBS JOURNAL, Issue 8 2002
Eleonora Cerasoli
Shikimate kinase was chosen as a convenient representative example of the subclass of ,/, proteins with which to examine the mechanism of protein folding. In this paper we report on the refolding of the enzyme after denaturation in urea. As shown by the changes in secondary and tertiary structure monitored by far UV circular dichroism (CD) and fluorescence, respectively, the enzyme was fully unfolded in 4 m urea. From an analysis of the unfolding curve in terms of the two-state model, the stability of the folded state could be estimated as 17 kJ·mol,1. Approximately 95% of the enzyme activity could be recovered on dilution of the urea from 4 to 0.36 m. The results of spectroscopic studies indicated that refolding occurred in at least four kinetic phases, the slowest of which (k = 0.009 s,1) corresponded with the regain of shikimate binding and of enzyme activity. The two most rapid phases were associated with a substantial increase in the binding of 8-anilino-1-naphthalenesulfonic acid with only modest changes in the far UV CD, indicating that a collapsed intermediate with only partial native secondary structure was formed rapidly. The relevance of the results to the folding of other ,/, domain proteins is discussed. [source]

Crystallization and preliminary X-ray diffraction analysis of shikimate kinase from Mycobacterium tuberculosis in complex with MgADP

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2001
Yijun Gu
Shikimate kinase (SK) from Mycobacterium tuberculosis (Mt) was overexpressed in Escherichia coli, purified and cocrystallized with MgADP in hanging drops using the vapor-diffusion procedure with PEG 4000 and 2-propanol as precipitants at pH 7.5. The crystal of MtSK,MgADP, which diffracted to 2.2,Å resolution, belonged to space group P3221 or P3121, with unit-cell parameters a = b = 64.01, c = 92.41,Å. There was one MtSK molecule in the asymmetric unit. Molecular-replacement trials with the crystal structure of SK from Erwinia chrysanthemi (PDB code 1shk) and adenylate kinase (PDB code 1ake) as search models were not successful. Heavy-atom derivative screening is in progress. [source]

Effects of the magnesium and chloride ions and shikimate on the structure of shikimate kinase from Mycobacterium tuberculosis

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2007
Marcio Vinicius Bertacine Dias
Bacteria, fungi and plants can convert carbohydrate and phosphoenolpyruvate into chorismate, which is the precursor of various aromatic compounds. The seven enzymes of the shikimate pathway are responsible for this conversion. Shikimate kinase (SK) is the fifth enzyme in this pathway and converts shikimate to shikimate-3-phosphate. In this work, the conformational changes that occur on binding of shikimate, magnesium and chloride ions to SK from Mycobacterium tuberculosis (MtSK) are described. It was observed that both ions and shikimate influence the conformation of residues of the active site of MtSK. Magnesium influences the conformation of the shikimate hydroxyl groups and the position of the side chains of some of the residues of the active site. Chloride seems to influence the affinity of ADP and its position in the active site and the opening length of the LID domain. Shikimate binding causes a closing of the LID domain and also seems to influence the crystallographic packing of SK. The results shown here could be useful for understanding the catalytic mechanism of SK and the role of ions in the activity of this protein. [source]

The refolding of type II shikimate kinase from Erwinia chrysanthemi after denaturation in urea

FEBS JOURNAL, Issue 8 2002
Eleonora Cerasoli
Shikimate kinase was chosen as a convenient representative example of the subclass of ,/, proteins with which to examine the mechanism of protein folding. In this paper we report on the refolding of the enzyme after denaturation in urea. As shown by the changes in secondary and tertiary structure monitored by far UV circular dichroism (CD) and fluorescence, respectively, the enzyme was fully unfolded in 4 m urea. From an analysis of the unfolding curve in terms of the two-state model, the stability of the folded state could be estimated as 17 kJ·mol,1. Approximately 95% of the enzyme activity could be recovered on dilution of the urea from 4 to 0.36 m. The results of spectroscopic studies indicated that refolding occurred in at least four kinetic phases, the slowest of which (k = 0.009 s,1) corresponded with the regain of shikimate binding and of enzyme activity. The two most rapid phases were associated with a substantial increase in the binding of 8-anilino-1-naphthalenesulfonic acid with only modest changes in the far UV CD, indicating that a collapsed intermediate with only partial native secondary structure was formed rapidly. The relevance of the results to the folding of other ,/, domain proteins is discussed. [source]

Crystallization and preliminary X-ray diffraction analysis of shikimate kinase from Mycobacterium tuberculosis in complex with MgADP

ACTA CRYSTALLOGRAPHICA SECTION D, Issue 12 2001
Yijun Gu
Shikimate kinase (SK) from Mycobacterium tuberculosis (Mt) was overexpressed in Escherichia coli, purified and cocrystallized with MgADP in hanging drops using the vapor-diffusion procedure with PEG 4000 and 2-propanol as precipitants at pH 7.5. The crystal of MtSK,MgADP, which diffracted to 2.2,Å resolution, belonged to space group P3221 or P3121, with unit-cell parameters a = b = 64.01, c = 92.41,Å. There was one MtSK molecule in the asymmetric unit. Molecular-replacement trials with the crystal structure of SK from Erwinia chrysanthemi (PDB code 1shk) and adenylate kinase (PDB code 1ake) as search models were not successful. Heavy-atom derivative screening is in progress. [source]

Effects of the magnesium and chloride ions and shikimate on the structure of shikimate kinase from Mycobacterium tuberculosis

ACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 1 2007
Marcio Vinicius Bertacine Dias
Bacteria, fungi and plants can convert carbohydrate and phosphoenolpyruvate into chorismate, which is the precursor of various aromatic compounds. The seven enzymes of the shikimate pathway are responsible for this conversion. Shikimate kinase (SK) is the fifth enzyme in this pathway and converts shikimate to shikimate-3-phosphate. In this work, the conformational changes that occur on binding of shikimate, magnesium and chloride ions to SK from Mycobacterium tuberculosis (MtSK) are described. It was observed that both ions and shikimate influence the conformation of residues of the active site of MtSK. Magnesium influences the conformation of the shikimate hydroxyl groups and the position of the side chains of some of the residues of the active site. Chloride seems to influence the affinity of ADP and its position in the active site and the opening length of the LID domain. Shikimate binding causes a closing of the LID domain and also seems to influence the crystallographic packing of SK. The results shown here could be useful for understanding the catalytic mechanism of SK and the role of ions in the activity of this protein. [source]