title: Elucidation of Mg²⁺ binding activity of adenylate kinase from Mycobacterium tuberculosis H₃₇Rv using fluorescence studies.
abstract: Adenylate kinase (AK) is a small ubiquitous enzyme that catalyzes the reversible transfer of the terminal phosphate group from adenine triphosphate (ATP): magnesium ion (Mg²⁺) to adenine monophosphate (AMP) to form two molecules of adenine diphosphate (ADP). AK thus maintains the homeostasis of adenine nucleotides in eukaryotes and prokaryotes. Because the [ATP]/[ADP] ratio is an important parameter in energy regulation in cells, Mg²⁺-activated AK has an important biological role, particularly in the case of bacteria, as imbalance in the ratio of [ATP]/[ADP] has been associated with alteration in its DNA supercoiling state. In the present study, magnesium-binding assays were carried out by systematically varying the concentrations of Mg²⁺, protein, AMP, ATP, and indicator in kinetic experiments. We report evidence that during magnesium-binding assay, the fluorescence level of the indicator "Mag-Indo-1" changes with protein concentration, suggesting that magnesium ions are binding to AK. The dual activity of AK both as nucleoside monophosphate and diphosphate kinases suggests that this enzyme may have a role in RNA and DNA biosynthesis in addition to its role in intracellular nucleotide metabolism. According to the proposed model, the magnesium-activated AK exhibits an increase in its forward reaction rate compared with the inactivated form. These findings imply that Mg²⁺ could be an important regulator in the energy signaling network in cells.
authors.affiliation: CSIR-Institute of Genomics and Integrative Biology, Delhi 110007, India. meena@igib.res.in
authors.forename: Laxman S
authors.initials: LS
authors.lastname: Meena
authors.forename: Sanjay R
authors.initials: SR
authors.lastname: Dhakate
authors.forename: Purushottam D
authors.initials: PD
authors.lastname: Sahare
mesh_terms: D000263:Adenylate Kinase / Q000378:metabolism*; D057075:Enzyme Assays / Q000379:methods*; D005453:Fluorescence; D008163:Luminescent Measurements / Q000379:methods*; D008274:Magnesium / Q000378:metabolism*; D009169:Mycobacterium tuberculosis / Q000201:enzymology*; D011485:Protein Binding; D013816:Thermodynamics
chemical_list: D000263:Adenylate Kinase; D008274:Magnesium