Protein N-myristoylation in Escherichia coli: Reconstitution of a eukaryotic protein modification in bacteria

Journal Article · Thu Feb 01 04:00:00 EST 1990 · Proceedings of the National Academy of Sciences of the United States of America; (USA)

DOI:https://doi.org/10.1073/pnas.87.4.1506· OSTI ID:6072973

Duronio, R J; Jackson-Machelski, E; Heuckeroth, R O; Gordon, J I ^[1]; Olins, P O; Devine, C S ^[2]; Yonemoto, W; Slice, L W; Taylor, S S ^[3]

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1. Washington Univ. School of Medicine, St. Louis, MO (USA)
2. Monsanto Co., St. Louis, MO (USA)
3. Univ. of California, San Diego (USA)

Protein N-myristoylation refers to the covalent attachment of a myristoyl group (C14:0), via amide linkage, to the NH{sub 2}-terminal glycine residue of certain cellular and viral proteins. Myristoyl-CoA:protein N-myristoyltransferase (NMT) catalyzes this cotranslational modification. The authors have developed a system for studying the substrate requirements and biological effects of protein N-myristoylation as well as NMT structure-activity relationships. Expression of the yeast NMT1 gene in Escherichia coli, a bacterium that has no endogenous NMT activity, results in production of the intact 53-kDa NMT polypeptide as well as a truncated polypeptide derived from proteolytic removal of its NH{sub 2}-terminal 39 amino acids. By using a dual plasmid system, N-myristoylation of a mammalian protein was reconstituted in E. coli by simultaneous expression of the yeast NMT1 gene and a murine cDNA encoding the catalytic (C) subunit of cAMP-dependent protein kinase (PK-A). A major advantage of the bacterial system over eukaryotic systems is the absence of endogenous NMT and substrates, providing a more straightforward way of preparing myristoylated, analog-substituted, and nonmyristoylated forms of a given protein for comparison of their structural and functional properties. The experimental system may prove useful for recapitulating other eukaryotic protein modifications in E. coli so that structure-activity relationships of modifying enzymes and their substrates can be more readily assessed.

• OSTI ID: 6072973
• Journal Information: Proceedings of the National Academy of Sciences of the United States of America; (USA), Journal Name: Proceedings of the National Academy of Sciences of the United States of America; (USA) Vol. 87:4; ISSN 0027-8424; ISSN PNASA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
BACTERIA
CARBOXYLIC ACIDS
DNA
ENZYME ACTIVITY
ENZYMES
ESCHERICHIA COLI
GLYCINE
HEXADECANOIC ACID
HYDROGEN COMPOUNDS
MICROORGANISMS
MOLECULAR STRUCTURE
MONOCARBOXYLIC ACIDS
NUCLEIC ACIDS
ORGANIC ACIDS
ORGANIC COMPOUNDS
PEPTIDES
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHOTRANSFERASES
POLYPEPTIDES
POST-TRANSLATION MODIFICATION
PROTEINS
RECOMBINANT DNA
STRUCTURE-ACTIVITY RELATIONSHIPS
SUBSTRATES
TETRADECANOIC ACID
TRANSFERASES
TRITIUM COMPOUNDS