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Published on: November 13, 2015
Comparative binding of lactate dehydrogenase to mitochondrial fractions
J Pruñonosa1, M L Sagristá, J Bozal
1Department of Biochemistry and Physiology, Faculty of Chemistry, University of Barcelona.
Abstract:
Beef liver mitochondrial fraction showed LDH activity (1.76 +/- 0.25 U/g pellet). Sixty seven% of the initial mitochondrial pellet LDH activity (almost M4 isoenzyme) was released when suspended in NaCl 0.15 M. When the washed particles were sonicated in a 0.15 M NaCl medium, the solubilized LDH activity (all five isoenzymes as cytosoluble fraction) was 5-fold higher than the initial pellet activity. The different isoenzymatic composition of intramitochondrial and externally bound forms of the enzyme should be taken into account when investigating the physiological role of intramitochondrial LDH. Beef liver cytosoluble LDH (very little content of M4 isoenzyme) showed no affinity for the beef liver mitochondrial fraction but purified M4-LDH isoenzyme was able to bind to the particulate fraction from the same source. This suggests an isoenzyme specificity for the interaction. The maximum amount of cytosoluble LDH bound to the mitochondrial fraction depends on the enzyme and the particulate fraction source. Therefore, binding capacity to the mitochondrial fraction depends not only on the net charge of LDH isoenzymes, which play a predominant role in the binding, but also on individual characteristics of the LDH isoenzymes and mitochondrial fractions from different sources. This suggests that electrostatic forces are not the only ones involved in the binding process.
Insights
Lactate dehydrogenase (LDH) activity in beef liver mitochondria was investigated. LDH isoenzymes exhibit specific binding to mitochondria, influenced by factors beyond net charge, suggesting complex interactions.
Area of Science:
- Biochemistry
- Cell Biology
- Enzymology
Background:
- Lactate dehydrogenase (LDH) is a crucial enzyme in cellular metabolism.
- Mitochondria play a vital role in cellular energy production and homeostasis.
- The localization and interaction of LDH isoenzymes with cellular compartments like mitochondria are not fully understood.
Purpose of the Study:
- To investigate the presence and activity of LDH within beef liver mitochondrial fractions.
- To determine the binding characteristics and specificity of different LDH isoenzymes to beef liver mitochondria.
- To elucidate the factors influencing LDH binding to mitochondrial fractions.
Main Methods:
- Isolation and characterization of beef liver mitochondrial fractions.
- Assay of LDH activity in mitochondrial pellets and soluble fractions.
- Investigation of LDH isoenzyme release and binding using varying salt concentrations and sonication.
- Analysis of LDH isoenzyme binding affinity to mitochondrial fractions.
Main Results:
- Beef liver mitochondrial fractions exhibited significant LDH activity, primarily the M4 isoenzyme.
- LDH activity was released from mitochondria in 0.15 M NaCl, with sonication solubilizing a 5-fold higher activity.
- Cytosolic LDH showed no affinity for mitochondria, while purified M4-LDH isoenzyme bound specifically, indicating isoenzyme specificity.
- Mitochondrial binding capacity depends on LDH isoenzyme characteristics and the source of the mitochondrial fraction, suggesting involvement of forces beyond net charge.
Conclusions:
- Intramitochondrial and externally bound LDH forms possess distinct isoenzymatic compositions.
- The interaction between LDH isoenzymes and beef liver mitochondria is specific and influenced by multiple factors.
- Electrostatic forces alone do not fully explain the binding of LDH isoenzymes to mitochondria.
