Characterization of random-sequence proteins displayed on the surface of Escherichia coli RNase HI.
In a previous study, random-sequence proteins of 120-130 amino acid residues were inserted into the surface loop region of the enzyme, Escherichia coli RNase HI [Doi et al. (1997) FEBS Lett. 402, 177-1801. Here we established that the RNase H activity of the insertion mutants is correlated with their secondary structure contents evaluated by circular dichroism measurement at 222 nm. The random-sequence insert of a mutant enzyme possessing relatively high RNase H activity was detached from the RNase HI scaffold, and its characterization indicated that the random-sequence protein maintains its secondary structure after separation from the scaffold. Thus, the structural features of random-sequence proteins were suggested to be monitored by measuring the activity of the scaffold enzyme into which these proteins have been inserted.
Doi
N
N
Mitsubishi Kasei Institute of Life Sciences, Machida, Tokyo, Japan.
Yomo
T
T
Itaya
M
M
Yanagawa
H
H
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics*; D017510:Protein Folding; D016914:Ribonuclease H / Q000737:chemistry* / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
D001426:Bacterial Proteins; D016914:Ribonuclease H; C078856:ribonuclease HI