Springer Nature is making SARS-CoV-2 and COVID-19 research free. View research | View latest news | Sign up for updates

Mechanisms of action of NIP71 on N-myristoyltransferase activity

  • 45 Accesses

  • 16 Citations


N-Myristoyl-CoA:protein N-myristoyltransferase (NMT) is the enzyme that catalyses the transfer of myristate from myristoyl-CoA to the N-terminal glycine of protein substrates. NMT was highly purified from bovine brain by procedures involving sequential column chromatography on DEAE-Sepharose CL-6B, phosphocellulose, hydroxylapatite, and mono S and mono Q f.p.l.c.. The highly purified NMT (termed NMT·II) possessed high specific activity with peptide substrates derived from the N-terminal sequences of the cAMP-dependent protein kinase and pp60src (29,800 and 47,600 pmol N-myristoylpeptide formed/min/mg, respectively), intermediate activity with a peptide based on the N-terminal sequence of a viral structural protein (μl) (M2; 17,300 pmol N-myristoylpeptide formed/min/mg) and very low activity with a peptide derived from the N-terminal sequence ofmyristoylatedalanine-richC-kinasesubstrate (MARCKS; 1500 pmol myristoylpeptide formed/min/mg). An NMT protein inhibitor (NIP71) isolated from the particulate fraction of bovine brain (King MJ and Sharma RK: Biochem J 291∶635-639, 1993) potently inhibited highly purified NMT activity (IC50 23.7 nM). A minor NMT activity (NMT·PU; 30% total NMT activity), which failed to bind to phosphocellulose, was insensitive to NIP71 inhibition. Inhibition of NMT was observed to be via mixed inhibition with respect to both the myristoyl-CoA and peptide substrates with NIP71 having an apparent higher affinity for NMT than the NMT·myristoyl·CoA complex. Inhibition by NIP71 at subsaturating concentrations of myristolyl-CoA and peptide resulted in a sigmoidal pattern of inhibition indicating that bovine brain possesses a potent and delicate on/off switch to control NMT activity.

This is a preview of subscription content, log in to check access.



N-myristoyl-CoA:protein N-myristoyltransferase


mono Q N-myristoyl-CoA:protein N-myristoyltransferase peak I


mono Q N-myristoyl-CoA:protein N-myristoyltransferase peak II


mono Q N-myristoyl-CoA:protein N-myristoyltransferase peak III

NIP71 :

71 kDa heat-stable N-myristoyltransferase inhibitor protein


  1. 1.

    Aitken A, Cohen P, Santikarn S, et al: Identification of the NH2-terminal blocking group of calcineurin B as myristic acid. FEBS Lett 150: 314–318, 1982

  2. 2.

    Carr SA, Biemann K, Shoji S, et al: n-Tetradecanoyl is the NH2-terminal blocking group of the catalytic subunit of cyclic AMP-dependent protein kinase from bovine cardiac muscle. Proc Natl Acad Sci USA 79: 6128–6131, 1982

  3. 3.

    Buss JE, Kamps, MP, Sefton BM: Myristic acid is attached to the transforming protein of Rous sarcoma virus during or immediately after synthesis and is present in both soluble and membrane-bound forms of the protein. Mol Cell Biol 4: 2697–2704, 1984

  4. 4.

    Bryant M, Ratner L: Myristoylation-dependent replication and assembly of human immunodeficiency virus 1. Proc Natl Acad Sci USA 87: 523–527, 1990

  5. 5.

    Weaver TA, Panganiban AT: N myristoylation of the spleen necrosis virus matrix protein is required for correct association of the Gag polyprotein with intracellular membranes and for particle formation. J Virol 64: 3995–4001, 1990

  6. 6.

    Towler D, Glaser L: Protein fatty acid acylation: Enzymatic synthesis of an N-myristoylglycyl peptide. Proc Natl Acad Sci USA 83: 2812–2816, 1986

  7. 7.

    Heuckeroth RO, Towler DA, Adams SP, et al.: ll-(Ethylthio)undecanoic acid. A myristic acid analogue of altered mydrophobicity which is functional for peptide N-myristoylation with wheat germ and yeast acyltransferase. J Biol Chem 263: 2127–2133, 1988

  8. 8.

    Deichaite I, Casson LP, Ling H-P, et al:In vitro synthesis of pp60src: Myristoylation in a cell free system. Mol Cell Biol 8: 4295–4301, 1988

  9. 9.

    Glover CJ, Goddard C, Felsted RL: N-Myristoylation of p60src. Identification of a myristoyl-CoA:glycylpeptide N-myristoyltransferase in rat tissue. Biochem J 250: 485–491, 1988

  10. 10.

    Towler DA, Adams SP, Eubanks SR, et al: Myristoyl CoA: protein N-myristoyltransferase activities from rat liver and yeast possess overlapping yet distinct peptide substrate specificities. J Biol Chem 263: 1784–1790, 1988

  11. 11.

    McIlhinney RAJ, McGlone K: Characterisation of a myristoyl CoA: glycylpeptide N-myristoyl transferase activity in rat brain: Subcellular and regional distribution. J Neurochem 54: 110–117, 1990

  12. 12.

    King MJ, Sharma RK: N-myristoyl transferase assay using phosphocellulose paper binding. Anal Biochem 199: 149–153, 1991

  13. 13.

    King MJ, Sharma RK: Demonstration of multiple forms of bovine brain myristoyl CoA:protein N-myristoyl transferase. Mol Cell Biochem 113: 77–81, 1992

  14. 14.

    Duronio RJ, Towler DA, Heuckeroth RO, et al.: Disruption of the yeast N-myristoyl transferase gene causes recessive lethality. Science 243: 796–800, 1989

  15. 15.

    Olson EN, Spizz G: Fatty acylation of cellular proteins. Temporal and subcellular differences between palmitate and myristate acylation. J Biol Chem 261: 2458–2466, 1986

  16. 16.

    da Silva AM, Klein C: A rapid posttranslational myristoylation of a 68-kDa protein inD. discoideum. J Cell Biol 111: 401–407, 1990

  17. 17.

    Aderem AA, Albert KA, Keum MM, et al: Stimulus-dependent myristoylation of a major substrate for protein kinase C. Nature 332: 362–364, 1988

  18. 18.

    King MJ, Sharma RK: Identification, purification and characterization of a membrane-associated N-myristoyltransferase inhibitor protein from bovine brain. Biochem J 291: 635–639, 1993

  19. 19.

    Towler DA, Stevens PA, Eubanks SR, et al: Purification and characterization of yeast myristoyl CoA:protein N-myristoyltransferase. Proc Natl Acad Sci USA 84: 2708–2712, 1987

  20. 20.

    Rudnick DA, McWherter CA, Rocque WJ, et al: Kinetic and structural evidence for a sequential ordered Bi Bi mechanism of catalysis bySaccharomyces cerevisiae myristoyl CoA:protein N-myristoyltransferase. J Biol Chem 266: 9732–9739, 1991

  21. 21.

    Duronio RJ, Jackson-Machelski E, Heuckeroth RO, et al: Protein N-myristoylation inEschrichia coli: Reconstitution of a eukaryotic protein modification in bacteria. Proc Natl Acad Sci USA 87: 1506–1510, 1990

  22. 22.

    McIlhinney RAJ, McGlone K, Willis AC: Purification and partial sequencing of myristoyl-CoA:protein N-myristoyltransferase from bovine brain. Biochem J 290: 405–410, 1993

  23. 23.

    Duronio RJ, Reed SI, Gordon JI: Mutations of human myristoyl-CoA: protein N-myristoyltransferase cause temperature-sensitive myristic acid auxotropy inSaccharomyces cerevisiae. Proc Natl Acad Sci USA 89: 4129–4133, 1992

  24. 24.

    Raju RVS, Kalra J, Sharma RK: Purification and properties of bovine spleen N-myristoyl CoA protein:N-myristoyltransferase. J Biol Chem 269: 12080–12083, 1994

  25. 25.

    Boutin JA, Ferry G, Ernould A-P, Maes P, Remond G, Vincent M: Myristoyl-CoA:protein N-myristoyltransferase activity in cancer cells. Purification and characterisation of a cytosolic isoform from the murine leukaemia cell line L1210. Eur J Biochem 214: 853–867 1993

  26. 26.

    Jayasuriya AK, Nibert ML, Fields BN: Complete nucleotide sequence of the M2 gene segment of retrovirus type 3 Dearing and analysis of its protein product μl. Virology 163: 591–602, 1988

  27. 27.

    Bradford MM: A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of proteindye binding. Anal Biochem 72: 248–254, 1976

  28. 28.

    Wray W, Boulikas T, Wray VP, et al: Silver staining of proteins in polyacrylamide gels. Anal Biochem 118: 197–203, 1981

  29. 29.

    Rudnick DA, Rocque WJ, McWherther CA, et al: Use of photoactivatable peptide substrates ofSaccharomyces cerevisiae myristoyl-CoA:protein N-myristoyltransferase (Nmtlp) to characterise a myristoyl-CoA-Nmtl p-peptide ternary complex and to provide evidence for an ordered reaction mechanism. Proc Natl Acad Sci USA 90: 1087–1091, 1993

  30. 30.

    Cornish-Bowden A: Fundamentals of enzyme kinetics. Butterworth & Co (Publishers) Ltd, London, 1979

  31. 31.

    Wu Z, Sharma RK, Wang JH: Catalytic and regulatory properties of calmodulin-stimulated phosphodiesterase isozymes. Adv Sec Mess Phosphopr Res 25: 29–41, 1992

  32. 32.

    Sharma RK, Kalra J: Characterisation of calmodulin-dependent cyclic nucleotide phosphodiesterase isozymes. Biochem J 298: 97–100 1994

  33. 33.

    Cohen P: The structure and regulation of protein phosphatases. Annu Rev Biochem 58: 453–508, 1989

  34. 34.

    King MJ, Pugazhenthi S, Khandelwal RL, et al.: Elevated N-myristoyl transferase activity is reversed by sodium orthovanadate in streptozotocin-induced diabetic rat. Bioch Biophys Acta 1165: 259–262, 1993

  35. 35.

    King MJ, Pugazhenthi S, Khandelwal RL, Sharma RK: Membrane-associated N-myristoyltransferase activity is reduced in obese (fa/fa) Zucker rat liver. Biochem Biophys Res Commun 196: 665–670, 1993

Download references

Author information

Correspondence to Rajendra K. Sharma.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

King, M.J., Sharma, R.K. Mechanisms of action of NIP71 on N-myristoyltransferase activity. Mol Cell Biochem 141, 79–86 (1994). https://doi.org/10.1007/BF00926170

Download citation

Key words

  • N-myristoyltransferase
  • NIP71
  • mixed inhibition
  • lipid