A nonconserved carboxy-terminal segment of GroEL contributes to reaction temperature.
The role of the C-terminal segment of the GroEL equatorial domain was analyzed. To understand the molecular basis for the different active temperatures of GroEL from three bacteria, we constructed a series of chimeric GroELs combining the C-terminal segment of the equatorial domain from one species with the remainder of GroEL from another. In each case, the foreign C-terminal segment substantially altered the active temperature range of the chimera. Substitution of L524 of Escherichia coli GroEL with the corresponding residue (isoleucine) from psychrophilic GroEL resulted in a GroE with approximately wild-type activity at 25 degrees C, but also at 10 degrees C, a temperature at which wild-type E. coli GroE is inactive. In a detailed look at the temperature dependence of the GroELs, normal E. coli GroEL and the L524I mutant became highly active above 14 degrees C and 12 degrees C respectively. Similar temperature dependences were observed in a surface plasmon resonance assay of GroES binding. These results suggested that the C-terminal segment of the GroEL equatorial domain has an important role in the temperature dependence of GroEL. Moreover, E. coli acquired the ability to grow at low temperature through the introduction of cold-adapted chimeric or L524I mutant groEL genes.
Nakamura
Takamichi
T
Laboratory of Molecular Microbiology, College of Agriculture, Ibaraki University, Ami 3-21-1, Inashiki, Ibaraki 300-0393, Japan.
Tanaka
Mitsuko
M
Maruyama
Akihiko
A
Higashi
Yowsuke
Y
Kurusu
Yasurou
Y
D000222:Adaptation, Physiological; D000595:Amino Acid Sequence; D018834:Chaperonin 60 / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D029968:Escherichia coli Proteins; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017510:Protein Folding; D011993:Recombinant Fusion Proteins; D013696:Temperature; D014162:Transfection
D018834:Chaperonin 60; D029968:Escherichia coli Proteins; D011993:Recombinant Fusion Proteins