CytochromecPeroxidase−CytochromecComplex:  Locating the Second Binding Domain on CytochromecPeroxidase with Site-Directed Mutagenesis†

Biochemistry ◽  
2000 ◽  
Vol 39 (33) ◽  
pp. 10132-10139 ◽  
Author(s):  
Valerie W. Leesch ◽  
Jordi Bujons ◽  
A. Grant Mauk ◽  
Brian M. Hoffman
Keyword(s):  
Binding Domain ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis

ΔLys120, a mutation which destabilizes the ribosome-binding domain of ribosomal protein L7/L12

1990 ◽  
Vol 68 (5) ◽  
pp. 832-838 ◽  
Author(s):  
Michael Laughrea ◽  
Eric Higgins
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Protein ◽  
Binding Domain ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Ribosome Binding ◽  
Mutant Genes

Five-residue-long deletions centered on A1a63, A1a75, and Glu118 of ribosomal protein L7/L12 gave low mutant yields (5% or less) when the mutant genes were cloned in phage M13mp18 and controlled by the L10 promotor. Deletions of Glu118–Lys120 or Lys120 (the COOH-terminus of L7/L12) gave higher mutant yields, up to 50% with L7/L12ΔLys120. L7/L12ΔLys120 was not preferentially found in the S100 and not preferentially removed by LiCl washing, but was preferentially extracted from 70S ribosomes in the presence of 28–35% ethanol in 0.25–0.5 M NH4Cl. It follows that ΔLys120 destabilizes the ribosome-binding domain of ribosomal protein L7/L12 in an ethanol-containing solvent, which raises the question whether Lys120 is part of the ribosome-binding domain of L7/L12 during some step of protein synthesis or whether it is essential to preserve the conformation of the physiological ribosome-binding domain under structurally stressful conditions.Key words: ribosome, ribosomal protein L7/L12, site-directed mutagenesis.

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