Thermus thermophilus L4 ribosomal protein: purification and sensitivity alteration against erythromycin of E. coli cells harboring a single amino acid mutant of TthL4 within its extended loop.
Protein L4 from the thermophilic bacterium Thermus thermophilus (TthL4) was heterologously overproduced in Escherichia coli cells and purified under native conditions by using ion exchange chromatography. Although it's known strong binding to RNA (23S rRNA as well as mRNA) the yield of the purified protein was 6 mg per 10 g of cells and it is similar to that referred for Thermotoga maritima L4 ribosomal protein. In addition, E. coli cells harboring the wild type Thermus thermophilus L4 (wtTthL4) ribosomal protein as well as its mutant having changed the highly conserved glutamic acid 56 by alanine (TthL4-Ala 56) were incorporated into E. coli ribosomes after transformation of the host cells with the recombined vector. The cells having incorporated the mutant TthL4-Ala56 are more sensitive against erythromycin related to that containing the wtTthL4 protein.The resistance to the drug indicates that the mutated amino acid Glu56 is probably critical for the local ribosomal conformation and that its mutation induces conformational disturbances that are "transferred" to the entrance of the major exit tunnel, the place where the drug does bind.
Leontiadou
F
F
Laboratory of Biochemistry, School of Chemistry, Aristotle University of Thessaloniki, Thessaloniki, Greece.
Tsagkalia
A
A
Choli-Papadopoulou
T
T
D000900:Anti-Bacterial Agents / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004352:Drug Resistance, Microbial; D004917:Erythromycin / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D017433:Protein Structure, Secondary; D012269:Ribosomal Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016963:Thermus thermophilus / Q000737:chemistry / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D012269:Ribosomal Proteins; C056368:ribosomal protein L4; D004917:Erythromycin