DOI: 10.1073/pnas.96.16.9003. Direct crosslinking of the antitumor antibiotic sparsomycin, and its derivatives, to A2602 in the peptidyl transferase center of 23S-like rRNA within ribosome-tRNA complexes

Direct crosslinking of the antitumor antibiotic sparsomycin, and its derivatives, to A2602 in the peptidyl transferase center of 23S-like rRNA within ribosome-tRNA complexes

10.1073/pnas.96.16.9003

Crossref journal-article

Proceedings of the National Academy of Sciences

Proceedings of the National Academy of Sciences (341)

Abstract

The antitumor antibiotic sparsomycin is a universal and potent inhibitor of peptide bond formation and selectively acts on several human tumors. It binds to the ribosome strongly, at an unknown site, in the presence of an N-blocked donor tRNA substrate, which it stabilizes on the ribosome. Its site of action was investigated by inducing a crosslink between sparsomycin and bacterial, archaeal, and eukaryotic ribosomes complexed with P-site-bound tRNA, on irradiating with low energy ultraviolet light (at 365 nm). The crosslink was localized exclusively to the universally conserved nucleotide A2602 within the peptidyl transferase loop region of 23S-like rRNA by using a combination of a primer extension approach, RNase H fragment analysis, and crosslinking with radioactive [ 125 I]phenol-alanine-sparsomycin. Crosslinking of several sparsomycin derivatives, modified near the sulfoxy group, implicated the modified uracil residue in the rRNA crosslink. The yield of the antibiotic crosslink was weak in the presence of deacylated tRNA and strong in the presence of an N-blocked P-site-bound tRNA, which, as was shown earlier, increases the accessibility of A2602 on the ribosome. We infer that both A2602 and its induced conformational switch are critically important both for the peptidyl transfer reaction and for antibiotic inhibition. This supposition is reinforced by the observation that other antibiotics that can prevent peptide bond formation in vitro inhibit, to different degrees, formation of the crosslink.

Bibliography

Porse, B. T., Kirillov, S. V., Awayez, M. J., Ottenheijm, H. C. J., & Garrett, R. A. (1999). Direct crosslinking of the antitumor antibiotic sparsomycin, and its derivatives, to A2602 in the peptidyl transferase center of 23S-like rRNA within ribosome-tRNA complexes. Proceedings of the National Academy of Sciences, 96(16), 9003â9008.

Authors 5

Bo T. Porse (first)
Stanislav V. Kirillov (additional)
Mariana J. Awayez (additional)
Harry C. J. Ottenheijm (additional)
Roger A. Garrett (additional)

References 24 Referenced 42

H C J Ottenheijm, L A G M van den Broek, J P G Ballesta, Z Zylicz Prog Med Chem 23, 220–268 (1986). / Prog Med Chem by Ottenheijm H C J (1986)
R A Garrett, C Rodriguez-Fonseca Ribosomal RNA: Structure, Evolution, Processing and Function in Protein Biosynthesis, eds R A Zimmermann, A E Dahlberg (CRC, Boca Raton, FL), pp. 327–355 (1995). / Ribosomal RNA: Structure, Evolution, Processing and Function in Protein Biosynthesis by Garrett R A (1995)
10.1016/S0079-6603(08)60809-0
10.1073/pnas.88.9.3725
10.1006/jmbi.1994.0135
10.1021/bi00104a011
10.1038/222356a0
10.1016/0092-8674(89)90128-1
10.1016/0300-9084(91)90157-V
10.1139/o95-095
10.1006/jmbi.1996.0454
10.1006/jmbi.1996.0455
V I Makhno, N N Peshin, Semenkov, P Yu., S V Kirillov Mol Biol 22, 528–536 (1988). / Mol Biol by Makhno V I (1988)
10.1006/jmbi.1998.2509
10.1002/j.1460-2075.1994.tb06432.x
10.1111/j.1432-1033.1978.tb20927.x
J Christiansen, J Egebjerg, N Larsen, R A Garrett Ribosomes and Protein Synthesis: A Practical Approach, ed G Spedding (Oxford Univ. Press, Oxford), pp. 229–252 (1990). / Ribosomes and Protein Synthesis: A Practical Approach by Christiansen J (1990)
10.1017/S135583829998202X
10.1016/0003-2697(82)90156-7
10.1021/jm00128a051
10.1007/978-3-642-81309-2
Kirillov S. V. Porse B. T. & Garrett R. A. (1999) RNA 5 in press. (10.1017/S1355838299990568)
10.1016/0300-9084(87)90207-0
E F Gale, E Cundliffe, P E Reynolds, R A Richmond, M J Waring The Molecular Basis of Antibiotic Action (Wiley, London), pp. 402–457 (1981). / The Molecular Basis of Antibiotic Action by Gale E F (1981)

Dates

Type	When
Created	23 years, 1 month ago (July 26, 2002, 10:38 a.m.)
Deposited	3 years, 4 months ago (April 13, 2022, 4:53 p.m.)
Indexed	11 months, 3 weeks ago (Sept. 5, 2024, 2:51 p.m.)
Issued	26 years ago (Aug. 3, 1999)
Published	26 years ago (Aug. 3, 1999)
Published Online	26 years ago (Aug. 3, 1999)
Published Print	26 years ago (Aug. 3, 1999)

Funders 0

None

BibTeX

@article{Porse_1999, title={Direct crosslinking of the antitumor antibiotic sparsomycin, and its derivatives, to A2602 in the peptidyl transferase center of 23S-like rRNA within ribosome-tRNA complexes}, volume={96}, ISSN={1091-6490}, url={http://dx.doi.org/10.1073/pnas.96.16.9003}, DOI={10.1073/pnas.96.16.9003}, number={16}, journal={Proceedings of the National Academy of Sciences}, publisher={Proceedings of the National Academy of Sciences}, author={Porse, Bo T. and Kirillov, Stanislav V. and Awayez, Mariana J. and Ottenheijm, Harry C. J. and Garrett, Roger A.}, year={1999}, month=aug, pages={9003–9008} }

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