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Mitochondrial arginine kinase from the heart of the horseshoe crabLimulus polyphemus

II. Catalytic properties and studies of potential functional coupling with oxidative phosphorylation

Journal of Comparative Physiology B volume 160, pages 459–468 (1990)Cite this article

Summary

Catalytic properties and potential functional coupling with the adenine nucleotide translocase (ANT) were studied in arginine kinase (AK) from the mitochondria of the heart of the horseshoe crab,Limulus polyphemus. Kinetic constants determined for cytoplasmic AK, AK in mitochondria, and mitochondrial AK tightly bound to membrane fragments were virtually identical, indicating that these enzyme fractions showed minimal differences in terms of catalytic properties. Kinetic constants were also evaluated in intact mitochondria in the presence and absence of oxidative phosphorylation. The Ka and Kia values for MgATP were identical. These data, coupled with the observation of identical rates of arginine phosphate production at a range of MgATP concentrations under both conditions, indicate that there is no functional coupling of mitochondrial AK with the ANT system. Additional studies showed that mitochondrial AK was more effective than exogenously supplied creatine kinase (CK) in stimulating mitochondrial respiration via phosphagen formation. AK, because of its position within the mitochondrial environment, appears to reduce diffusion problems for ADP by raising the local concentrations in the intermembrane space. Unlike vertebrate CK, where there is good evidence for functional coupling with the ANT system, we find no such functional coupling between AK and ANT inL. polyphemus mitochondria. However, mitochondrial AK probably plays a limited role in regulating respiratory control and ADP homeostasis in this system.

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Abbreviations

AK :

arginine kinase

AKc :

cytoplasmic arginine kinase

AKm :

mitochondrial arginine kinase

AK mf :

fast mitochondrial arginine kinase

AKms :

slow mitochondrial arginine kinase

ANT :

adenine nucleotide translocase

ArgP :

arginine phosphate

BSA :

bovine serum albumin

CK :

creatine kinase

CKm :

mitochondrial creatine kinase

CrP :

creatine phosphate

DTT :

dithiothreitol

G6PDH :

glucose-6-phosphate dehydrogenase

HEPES :

hydroxyethylpiperazine ethanesulfonic acid

HK :

hexokinase

K'eq :

equilibrium constant

K a :

ternary dissociation constant

K ia :

binary dissociation constant

KPi :

potassium phosphate

LDH :

lactate dehydrogenase

MAR :

mass action ratio

MgAc :

magnesium acetate

PEP :

phosphoenolpyruvate

PK :

pyruvate kinase

RCR :

respiratory control ratio

SD :

standard deviation

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Authors and Affiliations

  1. Department of Biological Science, Florida State University, 32306, Tallahassee, FL, USA

    C. Doumen & W. R. Ellington

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  1. C. Doumen
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  2. W. R. Ellington
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Doumen, C., Ellington, W.R. Mitochondrial arginine kinase from the heart of the horseshoe crabLimulus polyphemus . J Comp Physiol B 160, 459–468 (1990). https://doi.org/10.1007/BF01075678

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Key words

  • Arginine kinase
  • Horseshoe crab
  • Mitochondria
  • Compartmentation
  • ATP-buffering