Cloning and expression of the inositol monophosphatase gene from Methanococcus jannaschii and characterization of the enzyme
- Boston Coll., Chestnut Hill, MA (United States). Merkert Chemistry Center
Inositol monophosphatase plays a pivotal role in the biosynthesis of di-myo-inositol-1,1-phosphate, an osmolyte found in hyperthermophilic archaea. Given the sequence homology between the MJ109 gene product of Methanococcus jannaschii and human inositol monophosphatase, the MJ109 gene was cloned and expressed in Escherichia coli and examined for inositol monophosphatase activity. The purified MJ109 gene product showed inositol monophosphatase activity with kinetic parameters (K{sub m} = 0.091 {+-} 0.016 mM; V{sub max} = 9.3 {+-} 0.45 {micro}mol of P{sub i} min{sup {minus}1} mg of protein{sup {minus}1}) comparable to those of mammalian and E. coli enzymes. Its substrate specificity, Mg{sup 2+} requirement, Li{sup +} inhibition, subunit association (dimerization), and heat stability were studied and compared to those of other inositol monophosphatases. The lack of inhibition by low concentrations of Li{sup +} and high concentrations of Mg{sup 2+} and the high rates of hydrolysis of glucose-1-phosphate and p-nitrophenylphosphate are the most pronounced differences between the archaeal inositol monophosphatase and those from other sources. The possible causes of these kinetic differences are discussed, based on the active site sequence alignment between M. jannaschii and human inositol monophosphatase and the crystal structure of the mammalian enzyme.
- Sponsoring Organization:
- USDOE, Washington, DC (United States)
- DOE Contract Number:
- FG02-91ER20025
- OSTI ID:
- 642327
- Journal Information:
- Applied and Environmental Microbiology, Vol. 64, Issue 7; Other Information: PBD: Jul 1998
- Country of Publication:
- United States
- Language:
- English
Similar Records
Rv2131c gene product: An unconventional enzyme that is both inositol monophosphatase and fructose-1,6-bisphosphatase
Occurrence and role of di-myo-inositol-1,1[prime]-phosphate in methanococcus igneus