Mutagenesis of histidine 26 demonstrates the importance of loop-loop and loop-protein interactions for the function of iso-1-cytochrome c.
In yeast iso-1-cytochrome c, the side chain of histidine 26 (His26) attaches omega loop A to the main body of the protein by forming a hydrogen bond to the backbone atom carbonyl of glutamic acid 44. The His26 side chain also forms a stabilizing intra-loop interaction through a hydrogen bond to the backbone amide of asparagine 31. To investigate the importance of loop-protein attachment and intra-loop interactions to the structure and function of this protein, a series of site-directed and random-directed mutations were produced at His26. Yeast strains expressing these variant proteins were analyzed for their ability to grow on non-fermentable carbon sources and for their intracellular production of cytochrome c. While the data show that mutations at His26 lead to slightly decreased intracellular amounts of cytochrome c, the level of cytochrome c function is decreased more. The data suggest that cytochrome c reductase binding is affected more than cytochrome c oxidase or lactate dehydrogenase binding. We propose that mutations at this residue increase loop mobility, which, in turn, decreases the protein's ability to bind redox partners.
Fetrow
J S
JS
Department of Biological Sciences, Center for Biochemistry and Biophysical Chemistry, State University of New York, Albany 12222, USA. jacque@isadora.albany.edu
Dreher
U
U
Wiland
D J
DJ
Schaak
D L
DL
Boose
T L
TL
D000595:Amino Acid Sequence; D002455:Cell Division / Q000235:genetics; D003574:Cytochrome c Group / Q000737:chemistry*; D045304:Cytochromes c; D003576:Electron Transport Complex IV / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry; D006639:Histidine / Q000235:genetics*; D006860:Hydrogen Bonding; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed / Q000235:genetics; D009245:NADH Dehydrogenase / Q000378:metabolism; D011485:Protein Binding / Q000502:physiology; D017510:Protein Folding; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000737:chemistry*; D029701:Saccharomyces cerevisiae Proteins; D013053:Spectrophotometry; D013329:Structure-Activity Relationship
C014059:CYC1 protein, S cerevisiae; D003574:Cytochrome c Group; D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D006639:Histidine; D045304:Cytochromes c; D007770:L-Lactate Dehydrogenase; D009245:NADH Dehydrogenase; D003576:Electron Transport Complex IV