The consensus motif for N-myristoylation of plant proteins in a wheat germ cell-free translation system.
Protein N-myristoylation plays key roles in various cellular functions in eukaryotic organisms. To clarify the relationship between the efficiency of protein N-myristoylation and the amino acid sequence of the substrate in plants, we have applied a wheat germ cell-free translation system with high protein productivity to examine the N-myristoylation of various wild-type and mutant forms of Arabidopsis thaliana proteins. Evaluation of the relationship between removal of the initiating Met and subsequent N-myristoylation revealed that constructs containing Pro at position 3 do not undergo N-myristoylation, primarily because of an inhibitory effect of this amino acid on elimination of the initiating Met by methionyl aminopeptidase. Our analysis of the consensus sequence for N-myristoylation in plants focused on the variability of amino acids at positions 3, 6 and 7 of the motif. We found that not only Ser at position 6 but also Lys at position 7 affects the selectivity for the amino acid at position 3. The results of our analyses allowed us to identify several A. thaliana proteins as substrates for N-myristoylation that had previously been predicted not to be candidates for such modification with a prediction program. We have thus shown that a wheat germ cell-free system is a useful tool for plant N-myristoylome analysis. This in vitro approach will facilitate comprehensive determination of N-myristoylated proteins in plants.
Yamauchi
Seiji
S
Cell-Free Science and Technology Research Center and Venture Business Laboratory, Ehime University, Matsuyama, Japan.
Fusada
Naoki
N
Hayashi
Hidenori
H
Utsumi
Toshihiko
T
Uozumi
Nobuyuki
N
Endo
Yaeta
Y
Tozawa
Yuzuru
Y
D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D002474:Cell-Free System; D016384:Consensus Sequence; D019814:Myristic Acid / Q000378:metabolism*; D010936:Plant Extracts / Q000737:chemistry / Q000378:metabolism*; D014176:Protein Biosynthesis; D014908:Triticum
D029681:Arabidopsis Proteins; D010936:Plant Extracts; D019814:Myristic Acid