Ribosomal proteins L15 and L16 are mere late assembly proteins of the large ribosomal subunit. Analysis of an Escherichia coli mutant lacking L15.
The (minus L15) character from the Escherichia coli strain AM16.98 was transduced to an RNase-deficient strain in order to enable a reconstitution analysis. The following results were obtained. 1) The strain lacking L15 showed a 2-3-fold prolonged generation time and the 70 S ribosomes a reduced tendency toward dissociation. 2) Active particles could not be reconstituted unless L15 was added. Addition of L15 regained activity, even if L15 was added after the two-step procedure during a third incubation. However, a modification of the standard two-step reconstitution procedure (lowering NH4+ from 400 to 240 mM and the incubation temperature of the second step from 50 to 47 degrees C) yielded 100% active particles in the absence of L15. Active particles could be formed which even lacked L15, L16, and L30. Addition of either L15 or L16 accelerated the formation of active particles in the second step by a factor of five, and both proteins together by a factor of more than 20. 3) The activation energy of the rate-limiting step of the second incubation was surprisingly reduced for about 20 kcal/mol in the absence of L15, although the corresponding rates were two to five times slower. We conclude 1) that L15 and L16 are late assembly proteins which accelerate the formation of active particles during the late assembly but are neither needed for the early assembly nor essential for ribosomal functions; 2) that some routes of the late assembly (e.g. incorporation of L16) are changing their significance depending on the NH4+ concentration and the absence and presence of L15; and 3) that different reactions are rate limiting during the second step incubation in the presence and absence of L15, respectively, and that the corresponding reaction rates exhibit a different temperature dependence.
Max-Planck-Institut für Molekulare Genetik, Abteilung Wittmann, Berlin-Dahlem, West Germany.
F J
FJ
Franceschi
K H
KH
Nierhaus
D002872:Chromosome Deletion; D015180:Electrophoresis, Gel, Two-Dimensional; D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D007700:Kinetics; D008957:Models, Genetic; D009154:Mutation; D012269:Ribosomal Proteins / Q000235:genetics* / Q000302:isolation & purification; D012270:Ribosomes / Q000378:metabolism*; D014161:Transduction, Genetic
D012269:Ribosomal Proteins; C043649:ribosomal protein L15; C027937:ribosomal protein L16