T-jump Study of Calcium Binding Kinetics of Calcium Chelators

Cell Calcium. 1997 Oct;22(4):255-68. doi: 10.1016/s0143-4160(97)90064-6.

Abstract

Recent experimental studies have investigated the kinetic competition between calcium chelators and the secretion apparatus at a fast central synapse. Simultaneously, mathematical modelling studies indicate the importance of a quantitative knowledge of the binding kinetics of the chelators in studying fast physiological processes operating on a millisecond time scale. Using the temperature-jump relaxation method, I have studied the in vitro kinetics of Bis-Fura-2, Furaptra, Fluo-3, Calcium-Green-1, Calcium-Green-5N, Calcium-Orange-5N as well as EGTA, BAPTA and H-EDTA in conditions which are identical to those implemented in our patch clamp recordings, i.e. 100-140 mM CsCl, 20-40 mM Cs-HEPES, 8 mM NaCl, pH = 7.2 at 22 degrees C. The results can be summarized as follows: all fluorescent indicators have on rates in the range of 10(8)-10(9) M-1s-1. They differ significantly with respect to their off-rates from each other according to their affinities, ranging from 100 s-1 up to 26,000 s-1. BAPTA is kinetically almost indistinguishable from Fura-2. EGTA and H-EDTA have small binding rate constants for calcium in the range of 3 x 10(6) M-1s-1 since, at pH 7.20, protons need to be dissociated from the chelators before they can bind calcium ions.

MeSH terms

  • Aniline Compounds / metabolism
  • Calcium / metabolism*
  • Chelating Agents / metabolism*
  • Edetic Acid / metabolism
  • Egtazic Acid / analogs & derivatives
  • Egtazic Acid / metabolism
  • Fluorescent Dyes / metabolism*
  • Fura-2 / analogs & derivatives
  • Fura-2 / metabolism
  • Indicators and Reagents / metabolism
  • Organic Chemicals
  • Pentetic Acid / metabolism
  • Temperature
  • Xanthenes / metabolism

Substances

  • Aniline Compounds
  • Chelating Agents
  • Fluorescent Dyes
  • Indicators and Reagents
  • Organic Chemicals
  • Xanthenes
  • 2-(2-(5-carboxy)oxazole)-5-hydroxy-6-aminobenzofuran-N,N,O-triacetic acid
  • calcium green
  • calcium orange
  • Fluo-3
  • Egtazic Acid
  • Pentetic Acid
  • Edetic Acid
  • 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid
  • Calcium
  • Fura-2