Residues His-15 and Arg-17 of HPr Participate Differently in Catabolite Signal Processing via CcpA

• Published in: The Journal of biological chemistry Vol. 282; no. 2; pp. 1175 - 1182
• Main Authors: Horstmann, Nicola, Seidel, Gerald, Aung-Hilbrich, Lwin-Mar, Hillen, Wolfgang
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 12.01.2007; American Society for Biochemistry and Molecular Biology
• Subjects: Allosteric Regulation; Arginine - metabolism; Bacillus megaterium - genetics; Bacillus megaterium - metabolism; Bacillus subtilis - genetics; Bacillus subtilis - metabolism; Bacterial Proteins - chemistry; Bacterial Proteins - genetics; Bacterial Proteins - metabolism; Binding Sites; Carbon - metabolism; DNA-Binding Proteins - chemistry; DNA-Binding Proteins - genetics; DNA-Binding Proteins - metabolism; Escherichia coli - genetics; Escherichia coli - metabolism; Histidine - metabolism; Mutagenesis; Phosphoenolpyruvate Sugar Phosphotransferase System - chemistry; Phosphoenolpyruvate Sugar Phosphotransferase System - genetics; Phosphoenolpyruvate Sugar Phosphotransferase System - metabolism; Protein Structure, Tertiary; Repressor Proteins - chemistry; Repressor Proteins - genetics; Repressor Proteins - metabolism; Serine - metabolism; Signal Transduction - physiology; Surface Plasmon Resonance; Transcription Factors - chemistry; Transcription Factors - genetics; Transcription Factors - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M605854200

The carbon catabolite control protein A (CcpA) senses the physiological state of the cell by binding several effectors and responds with differential regulation of many genes in Bacilli. HPr-Ser46-P or Crh-Ser46-P interact with CcpA and stimulate binding to catabolite responsive elements. In addition, the glycolytic intermediates fructose 1,6-bisphosphate (FBP) and glucose 6-phosphate (Glc-6-P) stimulate HPr-Ser46-P but not Crh-Ser46-P binding to CcpA. The mechanisms by which coeffector binding to CcpA is linked to differential gene expression are unclear. To address this question we mutated residues participating in the interaction between HPr-Ser46-P or Crh-Ser46-P and CcpA and analyzed their effects on CcpA binding and stimulation of cre binding by surface plasmon resonance. The HPrH15A and CcpAD297A mutations do not affect complex formation but abolish FBP and Glc-6-P stimulation. Likewise, the CrhQ15H mutant becomes sensitive to these glycolytic intermediates. Hence, the contact of HPrHis-15 to Asp-297 in CcpA is a determinant for HPr specific FBP and Glc-6-P stimulation. The HPrR17A and -K mutants are both strongly impaired in stimulation of CcpA binding to cre, but only HPrR17A is defect in binding to CcpA indicating that these residues affect allostery of CcpA. Mutations of the residues of CcpA, which contact Arg-17 of HPr, exhibit differential effects on regulation of catabolic genes. Taken together, His-15 of HPr processes sensing information, while Arg-17 is involved in determining the genetic output.