Differential effects of replacing Escherichia coli ribosomal protein L27 with its homologue from Aquifex aeolicus.
The rpmA gene, which encodes 50S ribosomal subunit protein L27, was cloned from the extreme thermophile Aquifex aeolicus, and the protein was overexpressed and purified. Comparison of the A. aeolicus protein with its homologue from Escherichia coli by circular dichroism analysis and proton nuclear magnetic resonance spectroscopy showed that it readily adopts some structure in solution that is very stable, whereas the E. coli protein is unstructured under the same conditions. A mutant of E. coli that lacks L27 was found earlier to be impaired in the assembly and function of the 50S subunit; both defects could be corrected by expression of E. coli L27 from an extrachromosomal copy of the rpmA gene. When A. aeolicus L27 was expressed in the same mutant, an increase in the growth rate occurred and the "foreign" L27 protein was incorporated into E. coli ribosomes. However, the presence of A. aeolicus L27 did not promote 50S subunit assembly. Thus, while the A. aeolicus protein can apparently replace its E. coli homologue functionally in completed ribosomes, it does not assist in the assembly of E. coli ribosomes that otherwise lack L27. Possible explanations for this paradoxical behavior are discussed.
Maguire
B A
BA
Department of Biochemistry and Molecular Biology, University of Massachusetts, Amherst, Massachusetts 01003, USA.
Manuilov
A V
AV
Zimmermann
R A
RA
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D002942:Circular Dichroism; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D016946:Gram-Negative Aerobic Rods and Cocci / Q000235:genetics*; D009682:Magnetic Resonance Spectroscopy; D008565:Membrane Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D009154:Mutation; D012269:Ribosomal Proteins / Q000235:genetics* / Q000302:isolation & purification; D012270:Ribosomes / Q000737:chemistry; D016415:Sequence Alignment; D014169:Transformation, Bacterial
D001426:Bacterial Proteins; D008565:Membrane Proteins; D012269:Ribosomal Proteins; C409030:RpmA protein, Pseudomonas aeruginosa; C034661:ribosomal proteins L27