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Structural mapping of rabbit muscle phosphofructokinase. Distance between the adenosine cyclic 3',5'-phosphate binding site and a reactive sulfhydryl group.

Publication ,  Journal Article
Craig, DW; Hammes, GG
Published in: Biochemistry
January 22, 1980

The cAMP binding site of rabbit muscle phosphofructokinase has been labeled with the fluorescent molecule 5'-(p-fluorosulfonylbenzoyl)-2-aza-1,N6-ethenoadenosine. The most reactive sulfhydry- group of this modified enzyme, which is catalytically active, has been labeled with 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole and with N-]4-(dimethylamino)-3,5-dinitrophenyl]maleimide. The calculated distances between the cAMP binding site and the most reactive sulfhydryl group, as determined by resonance energy transfer measurements, are 31 and 26 A, respectively, for the two sulfhydryl group labels. Both steady-state and fluorescent -ifetime techniques were used to measure the energy transfer efficiencies in 50 mM potassium phosphate (pH 8.0) and 1 mM ethylenediaminetetraacetic acid, and a value of 2/3 was assumed for the donor-acceptor orientation factor. If the difference in calculated distances is attributed to a difference in the orientation factor for the two donor-acceptor ,airs, the actual distance between the cAMP ligand binding site and the most reactive sulfhydryl group on phosphofructokinase is shown to be 28 +/- 6 A.

Duke Scholars

Published In

Biochemistry

DOI

ISSN

0006-2960

Publication Date

January 22, 1980

Volume

19

Issue

2

Start / End Page

330 / 334

Location

United States

Related Subject Headings

  • Sulfhydryl Compounds
  • Spectrometry, Fluorescence
  • Rabbits
  • Protein Conformation
  • Protein Binding
  • Phosphofructokinase-1
  • Muscles
  • Kinetics
  • Fluorescent Dyes
  • Cyclic AMP
 

Citation

Journal cover image

Published In

Biochemistry

DOI

ISSN

0006-2960

Publication Date

January 22, 1980

Volume

19

Issue

2

Start / End Page

330 / 334

Location

United States

Related Subject Headings

  • Sulfhydryl Compounds
  • Spectrometry, Fluorescence
  • Rabbits
  • Protein Conformation
  • Protein Binding
  • Phosphofructokinase-1
  • Muscles
  • Kinetics
  • Fluorescent Dyes
  • Cyclic AMP