title: Genetic analysis of the nitrogen assimilation control protein from Klebsiella pneumoniae.
abstract: The nitrogen assimilation control protein (NAC) from Klebsiella pneumoniae is a typical LysR-type transcriptional regulator (LTTR) in many ways. However, the lack of a physiologically relevant coeffector for NAC and the fact that NAC can carry out many of its functions as a dimer make NAC unusual among the LTTRs. In the absence of a crystal structure for NAC, we analyzed the effects of amino acid substitutions with a variety of phenotypes in an attempt to identify functionally important features of NAC. A substitution that changed the glutamine at amino acid 29 to alanine (Q29A) resulted in a NAC that was seriously defective in binding to DNA. The H26D substitution resulted in a NAC that could bind and repress transcription but not activate transcription. The I71A substitution resulted in a NAC polypeptide that remained monomeric. NAC tetramers can bind to both long and shorter binding sites (like other LTTRs). However, the absence of a coeffector to induce the conformational change needed for the switch from the former to the latter raised a question. Are there two conformations of NAC, analogous to the other LTTRs? The G217R substitution resulted in a NAC that could bind to the longer sites but had difficulty in binding to the shorter sites, and the I222R and A230R substitutions resulted in a NAC that could bind to the shorter sites but had difficulty in binding properly to the longer sites. Thus, there appear to be two conformations of NAC that can freely interconvert in the absence of a coeffector.
authors.affiliation: Department of Molecular, Cellular, and Developmental Biology, the University of Michigan, Ann Arbor, MI 48109-1048, USA.
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Rosario
authors.forename: Brian K
authors.initials: BK
authors.lastname: Janes
authors.forename: Robert A
authors.initials: RA
authors.lastname: Bender
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002850:Chromatography, Gel; D018983:DNA Footprinting; D024202:Electrophoretic Mobility Shift Assay; D007711:Klebsiella pneumoniae / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D055503:Protein Multimerization; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D014157:Transcription Factors