The Dynamical Mechanism of Auto-Inhibition of AMP-Activated Protein Kinase

We use a novel normal mode analysis of an elastic network model drawn from configurations generated during microsecond all-atom molecular dynamics simulations to analyze the mechanism of auto-inhibition of AMP-activated protein kinase (AMPK). A recent X-ray and mutagenesis experiment (Chen, et al Nature 2009, 459, 1146) of the AMPK homolog S. Pombe sucrose non-fermenting 1 (SNF1) has proposed a new conformational switch model involving the movement of the kinase domain (KD) between an inactive unphosphorylated open state and an active or semi-active phosphorylated closed state, mediated by the autoinhibitory domain (AID), and a similar mutagenesis study showed that rat AMPK has the same auto-inhibition mechanism. However, there is no direct dynamical evidence to support this model and it is not clear whether other functionally important local structural components are equally inhibited. By using the same SNF1 KD-AID fragment as that used in experiment, we show that AID inhibits the catalytic function by restraining the KD into an unproductive open conformation, thereby limiting local structural rearrangements, while mutations that disrupt the interactions between the KD and AID allow for both the local structural rearrangement and global interlobe conformational transition. Our calculations further show that the AID also greatly impacts the structuring and mobility of the activation loop.

[1]  W. L. Jorgensen,et al.  Comparison of simple potential functions for simulating liquid water , 1983 .

[2]  Lawrence Shapiro,et al.  Crystal Structures of the Adenylate Sensor from Fission Yeast AMP-Activated Protein Kinase , 2007, Science.

[3]  Carsten Kutzner,et al.  GROMACS 4:  Algorithms for Highly Efficient, Load-Balanced, and Scalable Molecular Simulation. , 2008, Journal of chemical theory and computation.

[4]  Rodrigo Lopez,et al.  Clustal W and Clustal X version 2.0 , 2007, Bioinform..

[5]  C. Thornton,et al.  The regulation and function of mammalian AMPK‐related kinases , 2009, Acta physiologica.

[6]  T. Darden,et al.  A smooth particle mesh Ewald method , 1995 .

[7]  Zhi-Xin Wang,et al.  Structural insight into the autoinhibition mechanism of AMP-activated protein kinase , 2009, Nature.

[8]  Liang Tong,et al.  Crystal structure of the heterotrimer core of Saccharomyces cerevisiae AMPK homologue SNF1 , 2007, Nature.

[9]  B. Kemp,et al.  Functional Domains of the α1 Catalytic Subunit of the AMP-activated Protein Kinase* , 1998, The Journal of Biological Chemistry.

[10]  Liang Tong,et al.  An inhibited conformation for the protein kinase domain of the Saccharomyces cerevisiae AMPK homolog Snf1. , 2010, Acta crystallographica. Section F, Structural biology and crystallization communications.

[11]  H. Berendsen,et al.  Molecular dynamics with coupling to an external bath , 1984 .

[12]  Tara Davis,et al.  A conserved mechanism of autoinhibition for the AMPK kinase domain: ATP-binding site and catalytic loop refolding as a means of regulation. , 2010, Acta crystallographica. Section F, Structural biology and crystallization communications.

[13]  D. Hardie,et al.  AMP-activated protein kinase as a drug target. , 2007, Annual review of pharmacology and toxicology.

[14]  J. Kuriyan,et al.  The Conformational Plasticity of Protein Kinases , 2002, Cell.

[15]  D. Hardie,et al.  Development of protein kinase activators: AMPK as a target in metabolic disorders and cancer. , 2010, Biochimica et biophysica acta.

[16]  Chris Sander,et al.  The double cubic lattice method: Efficient approaches to numerical integration of surface area and volume and to dot surface contouring of molecular assemblies , 1995, J. Comput. Chem..

[17]  D. Hardie,et al.  AMP-activated/SNF1 protein kinases: conserved guardians of cellular energy , 2007, Nature Reviews Molecular Cell Biology.

[18]  B. Kemp,et al.  Structure and function of AMP‐activated protein kinase , 2009, Acta physiologica.

[19]  Teresa Head-Gordon,et al.  Instantaneous normal modes as an unforced reaction coordinate for protein conformational transitions. , 2010, Biophysical journal.

[20]  L. Young,et al.  A crystallized view of AMPK activation. , 2009, Cell metabolism.

[21]  J. Schlitter Estimation of absolute and relative entropies of macromolecules using the covariance matrix , 1993 .

[22]  R. Friesner,et al.  Evaluation and Reparametrization of the OPLS-AA Force Field for Proteins via Comparison with Accurate Quantum Chemical Calculations on Peptides† , 2001 .

[23]  Tao Pang,et al.  Conserved α-Helix Acts as Autoinhibitory Sequence in AMP-activated Protein Kinase α Subunits* , 2007, Journal of Biological Chemistry.

[24]  Berk Hess,et al.  LINCS: A linear constraint solver for molecular simulations , 1997, J. Comput. Chem..

[25]  Tirion,et al.  Large Amplitude Elastic Motions in Proteins from a Single-Parameter, Atomic Analysis. , 1996, Physical review letters.

[26]  Liang Tong,et al.  Biochemical and functional studies on the regulation of the Saccharomyces cerevisiae AMPK homolog SNF1. , 2010, Biochemical and Biophysical Research Communications - BBRC.

[27]  Kristiina Takkinen,et al.  Acta Crystallogr. Sect. F Struct. Biol. Cryst. Commun. , 2008 .

[28]  Cristian Micheletti,et al.  Small- and large-scale conformational changes of adenylate kinase: a molecular dynamics study of the subdomain motion and mechanics. , 2008, Biophysical journal.

[29]  David Carling,et al.  Structural basis for AMP binding to mammalian AMP-activated protein kinase , 2007, Nature.

[30]  Nathalie Reuter,et al.  Principal component and normal mode analysis of proteins; a quantitative comparison using the GroEL subunit , 2011, Proteins.

[31]  G. Schaftenaar,et al.  Molden: a pre- and post-processing program for molecular and electronic structures* , 2000, J. Comput. Aided Mol. Des..