Crystal structures of leucyl/phenylalanyl-tRNA-protein transferase and its complex with an aminoacyl-tRNA analog.
Eubacterial leucyl/phenylalanyl-tRNA protein transferase (L/F-transferase), encoded by the aat gene, conjugates leucine or phenylalanine to the N-terminal Arg or Lys residue of proteins, using Leu-tRNA(Leu) or Phe-tRNA(Phe) as a substrate. The resulting N-terminal Leu or Phe acts as a degradation signal for the ClpS-ClpAP-mediated N-end rule protein degradation pathway. Here, we present the crystal structures of Escherichia coli L/F-transferase and its complex with an aminoacyl-tRNA analog, puromycin. The C-terminal domain of L/F-transferase consists of the GCN5-related N-acetyltransferase fold, commonly observed in the acetyltransferase superfamily. The p-methoxybenzyl group of puromycin, corresponding to the side chain of Leu or Phe of Leu-tRNA(Leu) or Phe-tRNA(Phe), is accommodated in a highly hydrophobic pocket, with a shape and size suitable for hydrophobic amino-acid residues lacking a branched beta-carbon, such as leucine and phenylalanine. Structure-based mutagenesis of L/F-transferase revealed its substrate specificity. Furthermore, we present a model of the L/F-transferase complex with tRNA and substrate proteins bearing an N-terminal Arg or Lys.
Suto
Kyoko
K
Institute for Biological Resources and Functions, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba, Ibaraki 305-8566, Japan.
Shimizu
Yoshihiro
Y
Watanabe
Kazunori
K
Ueda
Takuya
T
Fukai
Shuya
S
Nureki
Osamu
O
Tomita
Kozo
K
D000595:Amino Acid Sequence; D019881:Aminoacyltransferases / Q000737:chemistry*; D001120:Arginine / Q000378:metabolism; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D008239:Lysine / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000378:metabolism; D017433:Protein Structure, Secondary; D011691:Puromycin / Q000737:chemistry*; D012346:RNA, Transfer, Amino Acyl / Q000737:chemistry*; D040681:Structural Homology, Protein
D029968:Escherichia coli Proteins; D050505:Mutant Proteins; D012346:RNA, Transfer, Amino Acyl; D011691:Puromycin; D001120:Arginine; D019881:Aminoacyltransferases; C021511:leucyltransferase; D008239:Lysine