ETD Collection

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Subject: search.filters.subject.Ribosomes

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    Preparation of ribsomal subunits by gel filtration
    (2016-07-20) Bhoolia, Deena
    An attempt was made to separate ribosomal subunits by gel filtl'ation on Trisacryl GF2000 and Sepharose 4B. Trisacryl GP~2000,a sYllthet'ic gol, , separated rat .ljver rlbosonal subunits orl 'f~'135,em column with a resolution of ""Or3, resulting 'in <'1" 60 rJ impm''ity 'of each of the \~ subunits. subunits" were not resolved. Sepharose 4131 an agarose based gol, separated " the subunits by adsorpttcn chromatography rathr.r than 'i>.V () Q ':) At 4°C, the 405 .subunf ts were eluted with a kd () ruO,:~9( but the GOrf' $ubud"its adsorbed to th~.'g,~Jl and were eluted I; /) when the temperature of "the column was increas~d to 250C..3SoC. (' ,. ('-. This edsorpt ion phenonenon seems to b{~ tl propert~ of a 11 agllrose -: \') ~\:; based gels studied here, includ'ing Sepharose 2.B and .seph'ato~eoBt arid is exc 1us iva to !,mamm'i'lan r ibosOIntrt subuni ts • Anal,ys1S 0'(" the , u subunits by in vitro r14C]polypheny1alanine sy'nthes'is showed OCI " ,,'c. /\ I 7', " diff'erence 'in the act lvtt ies of dbosoma'J ~ubunH~ p~epdrf#d by ;/'/ ' grndient centrifugation or by 5epharos,e cni"OmatogNPPY;' Analy~ii ~~of " (~ (( the subunits by ~crylamidec'ugarose coU)po5it~) gs1s resulted ,in the '/ resolution ,.:n"f subunits isolated fr'oill lower organisms in o II non-denaturlnq !Jcl systems and SI,lbut1its from m«mm~nan'tissue in II /'\';1 Ga'! f'iltrat'ion does tyffer a 5t!i:l;~'ble metrKld'¥or the pr~p;n~¥tiol1of <::) . I \) \\ I) ribosoma 1 'subunits I but only' if 'the act~orption JH'OrJ&~"t'Jo,s of ,) (,) ':' o ,1) ,{
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    Ribosomal RNA mutations to rifampicin resistance
    (2012-01-17) Macheke, Rulane Glenda
    In prokaryotes, transcription and translation are coupled and as a result, the beginning of the messenger RNA is translated by the ribosome while the 3' end is still synthesized. How exactly this occurs is still not clear. One possibility is that RNA polymerase and the ribosomes may be in physical contact with each other at some stage during gene expression or RNA polymerase has a binding site in the ribosomes. Mutational analysis is one method to explore how coordination between these two moieties occurs in bacteria. An Escherichia coli strain with all seven chromosomal ribosomal RNA operons deleted, replaced by a single rrnB plasmid-borne operon, was used to isolate ribosomal RNA mutants with increased rifampicin resistance, two of which were studied further. The altered rrnB operon in pGM1 was obtained by spontaneous whilst in pGM2 by EMS mutagenesis. The mutated rrnB operon in pGM1 conferred resistance to 25μg/ml of rifampicin while in pGM2 resistance of 30μg/ml was observed. A base substitution of T to A at position 355 of the 23S rRNA was detected in pGM1and no nucleotide change was detected in pGM2. The successful isolation of ribosomal RNA mutants with rifampicin resistance is consistent with the hypothesis of interaction between the RNA polymerase and the ribosomes and suggests the part of this interaction is with the large ribosomal subunit.