by: Robert J. Kosinski
Clemson University
Additional Background (and Some Corrections) on LDH
Fig. 1. A lactate dehydrogenase gel from Spring of 1998. The four lanes, right to left, are cow, goat, horse, and sheep.
This is presented to give you an idea of the expected results. You can
see the strong similarity between the goat and sheep lanes, with cow being
somewhat different and horse being very different. You should be able to explain
these results. However, the picture above does not mean you can ignore the
results you got this year. The gel above may be referred to in your Introduction
and certainly in your Discussion, but the only gel in your Results should
be the one you did yourself.
The M and H forms are specialized for work in different environments. Skeletal muscles are often anaerobic, and so the M form efficiently catalyzes the production of lactate from pyruvate (lactate fermentation, which regenerates NAD). However, the heart is hardly ever anaerobic, and so it would be a waste to turn valuable pyruvate into lactate there. The enzymatic activity of the H peptide is inhibited by pyruvate. This leaves the pyruvate alone so that it can enter the Krebs cycle and generate far more ATP for the heart muscle (Cahn et al., 1962). This difference can be seen in the distributions of the H and M monomers: in rat heart, 78% of the monomers are H; in rat leg muscles, only 11% of the monomers are H (Holbrook et al., 1975).
In a test tube, the H and the M forms will combine into tetramers randomly, depending on their relative abundance. However, in tissues, there are many molecules present that may not allow certain monomer combinations to occur (Glass and Doyle, 1972). Therefore, as development proceeds, different tissues get characteristic mixes of LDH tetramers (Holbrook et al., 1975).
For much more background on LDH structure and activity, check the Worthington Biochemical Corporation website on lactate dehydrogenase.
Fig. 2. Banding patterns produced by several animals in Cahn et al. (1962). In this case, the sera were all started at the center line, and bands could migrate either to the positive or negative electrode. HHHH will migrate the most strongly to the positive electrode and MMMM will tend to move to the negative electrode.
While it is certainly true that the tetramers of different animals migrate differently, it is less sure that phylogenetic relationships can be determined using LDH banding patterns. For example, rats and mice are obviously related, but their banding patterns in Fig. 2 are not very similar. Apparently, this problem does not exist in the hoofed mammals that we used.
One more recent study (Hiraoka et al., 1990) looked at evolutionary relationships using LDH in a different way. Rather than looking at electrophoretic mobility of the tetramers, they looked at the amino acid sequences of the monomers. This study showed two striking results for us:
Glass, R. and D. Doyle. 1972. Genetic control of lactate dehydrogenase expression in mammalian tissues. Science 176: 180-181.
Hiraoka, B. Y., F. S. Sharief, and Y. W. Yang. 1990. The cDNA and protein sequences of mouse lactate dehydrogenase B--molecular evolution of vertebrate lactate dehydrogenase genes A (muscle), B (heart) and C (testis). European J. Biochem. 189: 215-220.
Holbrook, J. J., A. Liljas, S. J. Steindel, and M. G. Rossman. 1975. Lactate dehydrogenase. pp. 191-292 in P. D. Boyer (Ed.), The Enzymes, Vol. XI, 3rd Ed. Academic Press, NY.
Li, S., W. Fitch, Y. Pan and F. Sharief. 1983. Evolutionary relationship of vertebrate lactate dehydrogenase isozyme A4 (muscle), B4 (heart) and C4 (testis). J. Biol. Chem. 258: 7029-7032.
Markert, C. L. and F. Moller. 1959. Multiple forms of enzymes: Tissue, ontogenetic, and species-specific patterns. Proc. Nat. Acad. Sci. US45: 753-763.