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Contribution of non-catalytic core residues to activity and regulation in protein kinase A.

Yang J, Kennedy EJ, Wu J, Deal MS, Pennypacker J, Ghosh G, Taylor SS
J Biol Chem 284 6241-8 (2009)
Cited: 31 times
EuropePMC logo PMID: 19122195

Abstract

Protein kinase A holoenzyme is comprised of two catalytic (C) and two regulatory (R) subunits which keep the enzyme in an inhibited state before activation by cyclic-AMP. The C-subunit folds into a conserved bi-lobal core flanked by N- and C-terminal tails. We report here characterization of a C-tail loss-of-function mutant, CF327A, and a related suppressor mutant, CF327A/K285P. Phe-327 is the only residue outside the kinase core that binds to the adenine ring of ATP, whereas Lys-285 is approximately 45 A away and lies in an AGC kinase-specific insert. The two mutations were previously identified from a yeast genetic screen, where the F327A mutation was unable to complement cell growth but mutation of K285P in the same allele rescued cell viability. We show that CF327A exhibits significant reduction in catalytic efficiency, which likely explains the observed loss-of-function phenotype. Interestingly, the additional K285P mutation does not restore kinase activity but reduces the inhibitory interaction of the double mutant with RII subunits. The additional K285P mutation, thus, helps to keep a low but uninhibited PKA activity that is sufficient for cell viability. The crystal structure of CF327A/K285P further reveals that recruitment of Phe-327 to the ATP binding pocket not only contributes to the hydrophobic pocket, as previously thought, but also recruits its flanking C-tail region to the kinase core, thereby concertedly positioning the glycine-rich loop and ATP for phosphoryl transfer. The study exemplifies two different ways for regulating cAMP-dependent protein kinase activity through non-conserved residues and sheds light on the structural and functional diversity of the kinase family.

Articles - 2qur mentioned but not cited (2)

  1. Contribution of non-catalytic core residues to activity and regulation in protein kinase A. Yang J, Kennedy EJ, Wu J, Deal MS, Pennypacker J, Ghosh G, Taylor SS. J Biol Chem 284 6241-6248 (2009)
  2. Crystal Structures Reveal Hidden Domain Mechanics in Protein Kinase A (PKA). Welsh CL, Conklin AE, Madan LK. Biology (Basel) 12 1370 (2023)


Reviews citing this publication (7)

  1. PKA: lessons learned after twenty years. Taylor SS, Zhang P, Steichen JM, Keshwani MM, Kornev AP. Biochim Biophys Acta 1834 1271-1278 (2013)
  2. Defining the conserved internal architecture of a protein kinase. Kornev AP, Taylor SS. Biochim Biophys Acta 1804 440-444 (2010)
  3. Evolution of the eukaryotic protein kinases as dynamic molecular switches. Taylor SS, Keshwani MM, Steichen JM, Kornev AP. Philos Trans R Soc Lond B Biol Sci 367 2517-2528 (2012)
  4. Solution NMR Spectroscopy for the Study of Enzyme Allostery. Lisi GP, Loria JP. Chem Rev 116 6323-6369 (2016)
  5. Allostery and binding cooperativity of the catalytic subunit of protein kinase A by NMR spectroscopy and molecular dynamics simulations. Masterson LR, Cembran A, Shi L, Veglia G. Adv Protein Chem Struct Biol 87 363-389 (2012)
  6. Exploring the Plasmodium falciparum cyclic-adenosine monophosphate (cAMP)-dependent protein kinase (PfPKA) as a therapeutic target. Haste NM, Talabani H, Doo A, Merckx A, Langsley G, Taylor SS. Microbes Infect 14 838-850 (2012)
  7. Phosphoproteomics Meets Chemical Genetics: Approaches for Global Mapping and Deciphering the Phosphoproteome. Jurcik J, Sivakova B, Cipakova I, Selicky T, Stupenova E, Jurcik M, Osadska M, Barath P, Cipak L. Int J Mol Sci 21 E7637 (2020)

Articles citing this publication (22)

  1. Crystal structure and allosteric activation of protein kinase C βII. Leonard TA, Różycki B, Saidi LF, Hummer G, Hurley JH. Cell 144 55-66 (2011)
  2. Turning enzymes ON with small molecules. Zorn JA, Wells JA. Nat Chem Biol 6 179-188 (2010)
  3. Dynamic architecture of a protein kinase. McClendon CL, Kornev AP, Gilson MK, Taylor SS. Proc Natl Acad Sci U S A 111 E4623-31 (2014)
  4. Structure and allostery of the PKA RIIβ tetrameric holoenzyme. Zhang P, Smith-Nguyen EV, Keshwani MM, Deal MS, Kornev AP, Taylor SS. Science 335 712-716 (2012)
  5. Localization and quaternary structure of the PKA RIβ holoenzyme. Ilouz R, Bubis J, Wu J, Yim YY, Deal MS, Kornev AP, Ma Y, Blumenthal DK, Taylor SS. Proc Natl Acad Sci U S A 109 12443-12448 (2012)
  6. Intramolecular C2 Domain-Mediated Autoinhibition of Protein Kinase C βII. Antal CE, Callender JA, Kornev AP, Taylor SS, Newton AC. Cell Rep 12 1252-1260 (2015)
  7. A transition path ensemble study reveals a linchpin role for Mg(2+) during rate-limiting ADP release from protein kinase A. Khavrutskii IV, Grant B, Taylor SS, McCammon JA. Biochemistry 48 11532-11545 (2009)
  8. Role of N-terminal myristylation in the structure and regulation of cAMP-dependent protein kinase. Bastidas AC, Deal MS, Steichen JM, Keshwani MM, Guo Y, Taylor SS. J Mol Biol 422 215-229 (2012)
  9. Cotranslational cis-phosphorylation of the COOH-terminal tail is a key priming step in the maturation of cAMP-dependent protein kinase. Keshwani MM, Klammt C, von Daake S, Ma Y, Kornev AP, Choe S, Insel PA, Taylor SS. Proc Natl Acad Sci U S A 109 E1221-9 (2012)
  10. A chimeric mechanism for polyvalent trans-phosphorylation of PKA by PDK1. Romano RA, Kannan N, Kornev AP, Allison CJ, Taylor SS. Protein Sci 18 1486-1497 (2009)
  11. A conserved Glu-Arg salt bridge connects coevolved motifs that define the eukaryotic protein kinase fold. Yang J, Wu J, Steichen JM, Kornev AP, Deal MS, Li S, Sankaran B, Woods VL, Taylor SS. J Mol Biol 415 666-679 (2012)
  12. Mutation of a kinase allosteric node uncouples dynamics linked to phosphotransfer. Ahuja LG, Kornev AP, McClendon CL, Veglia G, Taylor SS. Proc Natl Acad Sci U S A 114 E931-E940 (2017)
  13. Atomic Structure of GRK5 Reveals Distinct Structural Features Novel for G Protein-coupled Receptor Kinases. Komolov KE, Bhardwaj A, Benovic JL. J Biol Chem 290 20629-20647 (2015)
  14. Design and Profiling of a Subcellular Targeted Optogenetic cAMP-Dependent Protein Kinase. O'Banion CP, Priestman MA, Hughes RM, Herring LE, Capuzzi SJ, Lawrence DS. Cell Chem Biol 25 100-109.e8 (2018)
  15. Sensing domain dynamics in protein kinase A-I{alpha} complexes by solution X-ray scattering. Cheng CY, Yang J, Taylor SS, Blumenthal DK. J Biol Chem 284 35916-35925 (2009)
  16. Molecular features of product release for the PKA catalytic cycle. Bastidas AC, Wu J, Taylor SS. Biochemistry 54 2-10 (2015)
  17. Regulation of cAMP-dependent protein kinases: the human protein kinase X (PrKX) reveals the role of the catalytic subunit alphaH-alphaI loop. Diskar M, Zenn HM, Kaupisch A, Kaufholz M, Brockmeyer S, Sohmen D, Berrera M, Zaccolo M, Boshart M, Herberg FW, Prinz A. J Biol Chem 285 35910-35918 (2010)
  18. Zooming in on protons: Neutron structure of protein kinase A trapped in a product complex. Gerlits O, Weiss KL, Blakeley MP, Veglia G, Taylor SS, Kovalevsky A. Sci Adv 5 eaav0482 (2019)
  19. Structure of a PKA RIα Recurrent Acrodysostosis Mutant Explains Defective cAMP-Dependent Activation. Bruystens JG, Wu J, Fortezzo A, Del Rio J, Nielsen C, Blumenthal DK, Rock R, Stefan E, Taylor SS. J Mol Biol 428 4890-4904 (2016)
  20. Direct modulation of the protein kinase A catalytic subunit α by growth factor receptor tyrosine kinases. Caldwell GB, Howe AK, Nickl CK, Dostmann WR, Ballif BA, Deming PB. J Cell Biochem 113 39-48 (2012)
  21. Lysine acetylation modulates mouse sperm capacitation. Ritagliati C, Luque GM, Stival C, Baro Graf C, Buffone MG, Krapf D. Sci Rep 8 13334 (2018)
  22. Recognition of sites of functional specialisation in all known eukaryotic protein kinase families. Kalaivani R, Reema R, Srinivasan N. PLoS Comput Biol 14 e1005975 (2018)


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