A study of binding-solubilization of some glycolytic enzymes in striated muscle in situ

The solubilization of lactate dehydrogenase (LDH), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and alpha-glycerophosphate dehydrogenase (GAPDH) was studied in pressed muscle as a function of ionic strength and NADH concentration. The results indicate that these factors affect the binding-solubilization of LDH and GAPDH in a similar way to their effect in dilute homogenized tissue. Alpha-glycerophosphate dehydrogenase was included as a typical soluble enzyme, since we have been unable to demonstrate its binding to subcellular fractions under any conditions. As with homogenized tissue, LDH was less susceptible to solubilization by ionic strength than GAPDH. It was demonstrated that LDH isozymes richer in heart-type subunits were more easily removed from muscle by centrifugation-imbibition than isozymes richer in the muscle-type subunits. This was interpreted as indicating that binding of the enzyme to subcellular structures was a major factor in the restricted removal of these enzymes from muscle, since only the muscle-type subunit is capable of binding to subcellular particles. It was further demonstrated that LDH could be taken up into muscle tissue, depleted in the enzyme, against an apparent concentration gradient. This was also interpreted as binding of the enzyme to the particulate structure of the muscle. Furthermore, this uptake of LDH occurred under conditions of ionic strength (0.25) and pH (7.5) that would prevent binding of the enzyme to the particulate fraction of a dilute suspension of homogenized muscle tissue. Thus, physiological conditions of pH and ionic strength do not necessarily induce solubilization of chicken breast muscle LDH in situ. Data obtained with dilute tissue homogenates, therefore, may not necessarily be easily and safely extrapolated to conditions in situ.