DOI: 10.1111/j.1432-1033.1972.tb02066.x. Isolation of Active Ribosomal Subunits from Yeast

Isolation of Active Ribosomal Subunits from Yeast

10.1111/j.1432-1033.1972.tb02066.x

Crossref journal-article

Wiley

European Journal of Biochemistry (311)

Abstract

We have designed a method for the large scale isolation of pure and highly active subunits from yeast cytoplasmic ribosomes. The method consists of the incubation of yeast protoplasts in the presence of 1 mM NaN3. This causes the disappearence of polysomes and the formation of 80‐S ribosomes that can be dissociated in 0.5 M KCl. In this way over 90% of the ribosomes can be dissociated into subunits. The subunits are isolated by centrifugation in a B‐XV zonal rotor using an equivolumetric sucrose gradient. When this separation is performed at 5°C, dimerization of 40‐S particles to 65‐S particles occurs and no pure subunits can be obtained. At 10°C, there is no dimerization and the 60‐S + 40‐S subunits obtained are less than 1% cross‐contaminated. In poly(U)‐directed protein synthesis one subunit pair polymerizes phenylalanine residues at a rate of 1.7 residue per min. About 70% of the subunits were estimated to participate in this synthesis. Subunits prepared in the absence of dithiothreitol still bound phenylalanyl‐tRNA, but were inactive in phenylalanine polymerization. The 60‐S subunits prepared without dithiothreitol were as active as those prepared in the presence of dithiothreitol when their peptidyltransferase activity was tested.

Bibliography

van der Zeijst, B. A. M., Kool, A. J., & Bloemers, H. P. J. (1972). Isolation of Active Ribosomal Subunits from Yeast. European Journal of Biochemistry, 30(1), 15â25. Portico.

Authors 3

Ben A. M. van der Zeijst (first)
Addy J. Kool (additional)
Henri P. J. Bloemers (additional)

References 50 Referenced 49

10.1016/S0022-2836(63)80050-9
10.1073/pnas.60.2.569
10.1016/0022-2836(69)90084-9
10.1016/0022-2836(69)90085-0
10.1016/0006-291X(69)90892-4
10.1016/0022-2836(70)90042-2
10.1016/0006-291X(70)91086-7
10.1021/bi00811a008
10.1073/pnas.67.2.523 / Proc. Nat. Acad. Sci. U. S. A. by Leader D. P. (1970)
10.1016/0022-2836(69)90188-0
10.1073/pnas.68.2.390
10.1021/bi00790a018 / Biochemistry by Peterman M. L. (1971)
10.1016/0005-2787(71)90841-0
10.1021/bi00836a025 / Biochemistry by Dickhan S. R. (1969)
10.1016/0014-5793(69)80175-4
10.1016/0005-2787(71)90569-7
10.1111/j.1432-1033.1971.tb01501.x
10.1042/bj1240897 / Biochem. J. by Pratt H. (1971)
10.1101/SQB.1969.034.01.043
10.1016/0014-5793(70)80056-4
10.1016/0006-291X(71)90159-8
10.1016/0005-2787(71)90514-4
10.1016/0003-2697(71)90197-7
10.1016/0006-291X(71)90572-9
10.1128/jb.106.2.704-706.1971 / J. Bacteriol. by Etten J. L. (1971)
10.1021/bi00785a014
10.1016/S0021-9258(18)62379-8 / J. Biol. Chem. by Castles J. J. (1971)
10.1042/bj1240537
Engel C. J. M. Zeijst B. A. M.van der&Bloemers H. P. J.(1972) submitted for publication.
10.1016/S0021-9258(18)63264-8
10.1073/pnas.62.2.468
10.1128/JB.93.5.1662-1670.1967
Amesz W. J. C.&van derZeijst B. A. M.(1972)FEBS Lett. in press.
10.1016/0005-2787(68)90239-6
10.1038/215360a0
10.1016/0003-2697(68)90185-1
Reference deleted.
10.1016/0003-2697(70)90140-5
10.1016/0003-2697(71)90272-7
10.1111/j.1432-1033.1972.tb01933.x
10.1016/0005-2787(67)90677-6
{'key': 'e_1_2_3_43_2', 'first-page': '332', 'volume': '329', 'author': 'Merkenschlager M.', 'year': '1957', 'journal-title': 'Biochem. Z.'} / Biochem. Z. by Merkenschlager M. (1957)
{'key': 'e_1_2_3_44_2', 'first-page': '375', 'volume-title': 'Procedures in Nucleic Acid Research', 'author': 'Muench K. H.', 'year': '1966'} / Procedures in Nucleic Acid Research by Muench K. H. (1966)
Heller G.(1966)Dissertation Göttingen.
10.1016/0005-2787(66)90261-9
10.1042/bj0680653
10.1016/0006-291X(66)90586-9
10.1016/0022-2836(69)90075-8 / J. Mol. Biol. by Rychlík I. (1966)
10.1016/S0021-9258(18)62705-X
10.1038/newbio233257a0

Dates

Type	When
Created	20 years, 5 months ago (March 3, 2005, 9:57 a.m.)
Deposited	1 year, 9 months ago (Nov. 22, 2023, 4:47 p.m.)
Indexed	1 year, 7 months ago (Jan. 8, 2024, 7:59 p.m.)
Issued	52 years, 10 months ago (Oct. 1, 1972)
Published	52 years, 10 months ago (Oct. 1, 1972)
Published Online	20 years, 5 months ago (March 3, 2005)
Published Print	52 years, 10 months ago (Oct. 1, 1972)

Funders 0

None

BibTeX

@article{van_der_Zeijst_1972, title={Isolation of Active Ribosomal Subunits from Yeast}, volume={30}, ISSN={1432-1033}, url={http://dx.doi.org/10.1111/j.1432-1033.1972.tb02066.x}, DOI={10.1111/j.1432-1033.1972.tb02066.x}, number={1}, journal={European Journal of Biochemistry}, publisher={Wiley}, author={van der Zeijst, Ben A. M. and Kool, Addy J. and Bloemers, Henri P. J.}, year={1972}, month=oct, pages={15–25} }

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