Molecular Morphology of Ribosomes

Michalski, Chester J.; Sells, Bruce H.; Morison, Martin

doi:10.1111/j.1432-1033.1973.tb02706.x

Cited by 23 publications

(10 citation statements)

References 20 publications

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“…A probe such as lactoperoxidase-catalyzed iodination should have certain advantages over other systems. The iodination procedure results in little or no loss in biological activity or change in sedimentation behavior of the ribosomes [1,4,5]. Lactoperoxidase, unlike smaller-molecular-weight probes, should not penetrate the structure of the ribosome.…”

Section: Discussionmentioning

confidence: 99%

Molecular Morphology of Ribosomes

Michalski

Sells

1975

European Journal of Biochemistry

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Using either soluble or solid-state lactoperoxidase, a comparison was made between the enzymic iodination of ribosomal proteins iodinated as 3 0 4 and 50-S subunits or as 7 0 4 monosomes. Proteins S7, S11 and S12 of the 30-S subunit and proteins L2, L11, L26 and L28 of the 50-S subunit were labelled to a greater extent in isolated particles than in the 70-S ribosome. In contrast, proteins S4, S6, S19 and S20 were labelled to a lesser extent in the isolated subunit. No significant differences were observed in the iodination patterns of ribosomes iodinated in the presence of soluble lactoperoxidase and those iodinated in the presence of lactoperoxidase bound to Sepharose 4B. It is suggested that the 30-S subunit undergoes a conformational change during its association with the 50-S subunit to form a 70-S monosome. Implications from results obtained with solid-state lactoperoxidase-catalyzed iodination of ribosomal proteins are also discussed.Enzymic iodination of ribosomal proteins has been employed as a probe for the identification of surface proteins and detection of conformational changes in ribosome structure [l -41. Although a greater majority of 30-S proteins are extensively modified as compared to those of the 5 0 4 subunit, these studies reveal that essentially all of the 30-S and 50-S proteins become labelled, to some extent, with iodide. To eliminate the possibility that the accessibility of the individual ribosomal proteins to iodination is related to the ability of the lactoperoxidase to penetrate the structure of the ribosome, use was made of lactoperoxidase immobilized by covalent coupling to Sepharose 4B. Employing both soluble and immobilized lactoperoxidase, we have compared the ability of these two preparations to iodinate ribosomal proteins of isolated 3 0 3 and 5 0 4 subunits as well as subunit proteins derived from iodinated 70-S monosomes. Differences in iodination patterns between ribosomal proteins derived from isolated subunits and from 70-S ribosomes are also discussed with respect to structural alterations. MATERIALS AND METHODSBacterial strains, ribosome preparations, soluble lactoperoxidase-catalyzed iodinations and extraction of ribosomal proteins were as previously described [I, 51. 70-S monosomes were maintained at 10 mM Mg + throughout these experiments. Preparation of Solid-state LactoperoxidaseSolid-state lactoperoxidase was prepared according to the method described by David [6]. Sepharose 4B (5.0ml packed volume) was mixed with an equal volume of deionized water and activated with 2.5 g of cyanogen bromide; the reaction mixture, maintained at 20 "C, was immediately brought to pH 11 .O by the addition of 2 M NaOH. This pH was maintained by the further addition of 2 M NaOH until completion of the reaction as indicated by stabilization of pH. The temperature then was lowered to 4°C and the reaction mixture transferred to a Buchner funnel and washed under suction with phosphatebuffered saline. Washed Sepharose beads were then combined with purified lactoperoxidase [8] at a concentr...

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Section: Discussionmentioning

confidence: 99%

Molecular Morphology of Ribosomes

Michalski

Sells

1975

European Journal of Biochemistry

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“…Growth conditions of E. coli MRE600, labelling of r-proteins with [3H] tyrosine, ribosome preparations and extraction of r-proteins for gel electrophoresis were as previously described [ 19,21,23] . Lactoperoxidase obtained from Sigma Chemical Co. was further purified according to the method of Morrison and Hultquist [24].…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Molecular morphology of ribosomes: Effect of chain initiation factor 3 on 30 S subunit conformation

1976

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“…Cells of E. eoli MRE 600 were grown at 37°C as previously described (7,8 ) in the presence of tritiated tyrosine (5 pCiimL) ( I Ci = 37 GBq) for three generations to label celi proteins uniformly with tritium. Cell-free extracts following removal of cellular debris (4) were centrifuged at 48 000 rpm for 2.5 h in a Beckman type 50.2 Ti rotor and the resulting ribosome pellets were washed twice with 1 M NH,Cl, 10 mLW Tris-HCi, pH '7.4, 1 mLW magnesium acetate, and 6 mM 2-mercaptoethanoI.…”

Section: Preparation Sf Ribosomal Subunitsmentioning

confidence: 99%

Influence of magnesium and polyamines on the reactivity of individual ribosomal subunit proteins to lactoperoxidase-catalyzed iodination

Michalski¹,

Boyle²,

Sells³

1979

Can. J. Biochem.

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30S and 50S subunits, in the presence of either 20 mM Mg2+ or 6 mM Mg2+ and 5mM spermidine plus 25 mM putrescine, were observed to completely associate to form 70S monosomes as monitored by sucrose gradient sedimentation. Subunits maintained under the above ionic conditions were compared with 30S and 50S particles at low (6 mM) magnesium concentration with respect to the reactivity of individual ribosomal proteins to lactoperoxidase-catalyzed iodination. Altered reactivity to enzymatic iodination of ribosomal proteins S4, S9, S10, S14, S17, S19, and S20 in the small subunit of ribosomal proteins, L2, L9, L11, L27, and L30 in the large subunit following incubation with high magnesium or magnesium and polyamines suggests that a conformation change in both subunits accompanies the formation of 70S monosomes. The results further demonstrate that the effect of Mg2+ on subunit conformation is mimicked when polyamines are substituted for magnesium necessary for subunit association.

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Molecular Morphology of Ribosomes

Cited by 23 publications

References 20 publications

Molecular Morphology of Ribosomes

Molecular Morphology of Ribosomes

Molecular morphology of ribosomes: Effect of chain initiation factor 3 on 30 S subunit conformation

Influence of magnesium and polyamines on the reactivity of individual ribosomal subunit proteins to lactoperoxidase-catalyzed iodination

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