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J. Biochem, 1982, Vol. 91, No. 4 1241-1248
© 1982 Japanese Biochemical Society

research-article

The Nucleotide Sequence and Codon Recognition of Proline tRNA fromTorulopsis utilis

Kazuko OGAWA, Makoto KAWAKAMI, Yoshiko SHIMIZU1 and Shosuke TAKEMURA

Institute of Molecular Biology, School of Science, Nagoya University Chikusa-ku, Nagoya, Aichi 464

Proline tRNA from Torulopsis (Candida) utilis was purified by chromatography on columns of DEAE-Sephadex A-50 at pH 7.6 and 4.0, DEAE-Sephadex A-50 at pH 6.0, and RPC-5 at pH 4.0. This tRNA was sequenced by combining conventional chromatographic methods with rapid read-out chemical and enzymatic gel sequencing methods. The tRNA consists of 75 nucleotides and has the following sequence: pG-G-C-Cm-G-C-G-U-m1-G-G-U-C-{Psi}-A-G-D-G-G-D-A-U-A-C-U-C-U-C-G-C-U-U-N-G-G-m1-G-{Psi}-G-{Psi}-G-A-G-U-G-7-D-,m5-m5C-A-G-G-G-T-{Psi}-C-A-m1A-U-U-C-C-C-U-G-C-U-C-G-G-C-C-C-C-C-A. An unknown nucleotide in the first position of the anticodon was shown to be an uridine derivative by UV-spectrum, mobility on thin-layer chromatogram, and preliminary measurement of NMR spectrum. The specificity of this tRNA for codon recognition was studied by the ribo-somal binding technique. Three out of four proline codons were recognized: CCA and CCU preferentially, and CCC weakly. Unexpectedly, CCG failed to bind this tRNA to ribosomes.

1Present address: Department of Cellular and Developmental Biology, University of Arizona, Tucson, Arizona 85721, U.S.A.


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