Kinetic and thermodynamic analysis of RNA-protein interactions in the RNase P holoenzyme from Escherichia coli

Simon Talbot, S Altman

Research output: Contribution to journalArticlepeer-review

Abstract

A gel retardation assay has been used to examine the kinetic and equilibrium properties of the interaction between C5 protein and M1 RNA in the formation of the ribonuclease P holoenzyme from Escherichia coli. The interaction is relatively insensitive to the identity of the monovalent anions present and to pH in the range 7.0-9.0, but it has a more critical requirement for specific monovalent and divalent cations: NH4+, K+, Mg2+, Ca2+, and Mn2+ all promote efficient formation of the complex. A positive delta S (+6.4 cal mol-1 deg-1) and a negative delta H (-11.3 kcal mol-1) combine to give a delta G equal to -13.3 kcal mol-1 at 37 degrees C in 0.42 M salt. The binding reaction is sensitive to the concentration of monovalent and divalent cations, with the affinity increasing with increasing ionic strength (delta log Ka/delta log [NH4+] = +2.7 +/- 0.1). The dependence of Kd on the ionic strength and the positive delta S suggests that hydrophobic and stacking interactions contribute significantly to the formation of the RNase P holoenzyme.
Original languageEnglish
Pages (from-to)1406-11
Number of pages6
JournalBiochemistry
Volume33
Issue number6
Publication statusPublished - 1994

Keywords

  • Amino Acid Sequence
  • Bacterial Proteins
  • Cations, Divalent
  • Endoribonucleases
  • Escherichia coli
  • Escherichia coli Proteins
  • Hydrogen-Ion Concentration
  • Kinetics
  • Molecular Sequence Data
  • Osmolar Concentration
  • RNA, Catalytic
  • Ribonuclease P
  • Sequence Homology, Amino Acid
  • Solutions
  • Thermodynamics

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