title: Deciphering the catalysis-associated conformational changes of human adenylate kinase 1 with single-molecule spectroscopy.
abstract: Human adenylate kinase isoenzyme 1 (AK1) is the key enzyme in maintaining the cellular energy homeostasis. The catalysis-associated conformational changes of AK1 involve large-amplitude rearrangements. To decipher the conformational changes of AK1 at the single-molecule level, we tagged AK1 with two identical fluorophores, one near the substrate-binding site and the other at the boundary of the core domain. We found that magnesium ion binding to AK1 increases the structural heterogeneity of AK1, whereas ADP binding reduces the structural heterogeneity. We exploited the hidden Markov model to extract the underlying catalysis-associated conformational dynamics and determined thermodynamic parameters of the multiple catalytic pathways. The third-order correlation difference calculated from photon fluctuation traces reveals the irreversible nature of the conformational motions, suggesting that single-molecule AK1 is in a nonequilibrium steady state. This discovery offers a fresh viewpoint to look into the molecular mechanisms of cellular biochemistry.
authors.affiliation: Department of Photonics, Chiao Tung University , Hsinchu, Taiwan.
authors.forename: Chien Y
authors.initials: CY
authors.lastname: Lin
authors.forename: Jung Y
authors.initials: JY
authors.lastname: Huang
authors.forename: Leu-Wei
authors.initials: LW
authors.lastname: Lo
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry / Q000378:metabolism; D000263:Adenylate Kinase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001665:Binding Sites; D055162:Biocatalysis; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D008390:Markov Chains; D046210:Microfluidic Analytical Techniques; D056004:Molecular Dynamics Simulation; D017785:Photons; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D013379:Substrate Specificity; D013816:Thermodynamics
chemical_list: D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D000263:Adenylate Kinase; C110939:adenylate kinase 1