Functional domains of the InsA protein of IS2.
The InsA protein is a transcriptional regulator. It binds to the promoter region of insA and insAB'. To understand the molecular mechanism for the interaction between InsA and its binding sequence, the functional domains of InsA were identified. The glutaraldehyde cross-linking method and the two-hybrid expression system were used to study the protein-protein interaction of InsA. The results of these experiments showed that InsA forms homodimers. Deletion of the last 44 amino acid residues at its C terminus, but not the first 12 or 57 residues at the N terminus, abolished the ability of InsA to form homodimers, indicating that the protein-protein interaction domain of InsA is located at its C terminus. Gel retardation assays revealed that deletion of the last 29 amino acid residues at its C terminus had no effect on the DNA binding ability of InsA. In contrast, deletion of the first N-terminal 12 residues abolished the DNA binding capability of InsA. These results indicate that the DNA binding domain of InsA is located at its N terminus.
Lei
G S
GS
Department of Microbiology and Graduate Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan, Republic of China.
Hu
S T
ST
D001483:Base Sequence; D003001:Cloning, Molecular; D003432:Cross-Linking Reagents; D004251:DNA Transposable Elements; D004269:DNA, Bacterial / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D004926:Escherichia coli / Q000235:genetics; D029968:Escherichia coli Proteins; D005822:Genetic Vectors; D005976:Glutaral; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics
D003432:Cross-Linking Reagents; D004251:DNA Transposable Elements; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D029968:Escherichia coli Proteins; D012097:Repressor Proteins; C064548:insA1 protein, E coli; D005976:Glutaral