Lactate dehydrogenase patterns

Fig. 27. Lactate dehydrogenase patterns obtained by disc electrophoretic treatment of the enzyme from highly purified fractions of rat liver parenchymal a and Kupffer cells b. (After Berg and Blix, 1973)...

Fig. 3. Lactate dehydrogenase patterns on cellulose acetate strips of diaphragm and of in vitro cultured cells. The gels on the left show the five bands of mouse (M) and rat (R) diaphragm. On the right, patterns exhibited by homogenates of mouse and rat parents (MP and RP) and of hybrid Rj-aD (H).

The shift in pattern of protein synthesis during anaerobiosis has been observed in root tissue of many other plant species including rice, sorghum, barley, pea, and carrot (see Sachs Ho, 1986). In anaerobically treated barley aleurone cells, lactate dehydrogenase (LDH) activity increases (Hanson Jacobsen, 1984) as does enzyme activity and mRNA levels for ADH (Hanson, Jacobsen Zwar, 1984). 

Figure 7-11. Normal and pathologic patterns of lactate dehydrogenase (LDH) isozymes in human serum. LDH isozymes of serum were separated by electrophoresis and visualized using the coupled reaction scheme shown on the left. (NBT, nitroblue tetrazolium PMS, phenazine methylsulfate). At right is shown the stained electropherogram. Pattern A is serum from a patient with a myocardial infarct B is normal serum and C is serum from a patient with liver disease. Arabic numerals denote specific LDH isozymes.

The remaining three antiparallel /3 structures form a miscellaneous category (see Fig. 84). Lactate dehydrogenase d2 and gene 5 protein each has several two-stranded antiparallel j8 ribbons, but they do not coalesce into any readily described overall pattern. The N-terminal domain of tomato bushy stunt virus protein has a unique /3 structure in which equivalent pieces of chain from three different subunits wrap around a 3-fold axis to form what has been called a /3 annulus (Harrison et ah, 1978). Each of the three chains contributes a short strand segment to each of three three-stranded, interlocking /3 sheets. This domain provides one of the subunit contacts that hold the virus... 

An example of an enzyme which has different isoenzyme forms is lactate dehydrogenase (LDH) which catalyzes the reversible conversion of pyruvate into lactate in the presence of the coenzyme NADH (see above). LDH is a tetramer of two different types of subunits, called H and M, which have small differences in amino acid sequence. The two subunits can combine randomly with each other, forming five isoenzymes that have the compositions H4, H3M, H2M2, HM3 and M4. The five isoenzymes can be resolved electrophoretically (see Topic B8). M subunits predominate in skeletal muscle and liver, whereas H subunits predominate in the heart. H4 and H3M isoenzymes are found predominantly in the heart and red blood cells H2M2 is found predominantly in the brain and kidney while HM3 and M4 are found predominantly in the liver and skeletal muscle. Thus, the isoenzyme pattern is characteristic of a particular tissue, a factor which is of immense diagnostic importance in medicine. Myocardial infarction, infectious hepatitis and muscle diseases involve cell death of the affected tissue, with release of the cell contents into the blood. As LDH is a soluble, cytosolic protein it is readily released in these conditions. Under normal circumstances there is little LDH in the blood. Therefore the pattern of LDH isoenzymes in the blood is indicative of the tissue that released the isoenzymes and so can be used to diagnose a condition, such as a myocardial infarction, and to monitor the progress of treatment. 

In one study of patients receiving heparin, the isoenzyme pattern of lactate dehydrogenase was studied all had rises in the hepatic form of the enzyme, suggesting hepatocellular damage as the most likely source (60). 

FIGURE 1.14 Seen here is the hk0 zone diffraction pattern from a crystal of M4 dogfish lactate dehydrogenase obtained using a precession camera. It is based on a tetragonal crystal system and, therefore, exhibits a fourfold axis of symmetry. The hole at center represents the point where the primary X-ray beam would strike the film (but is blocked by a circular beamstop). Note the very predictable positions of the diffraction intensities. All the intensities, or reflections, fall at regular intervals on an orthogonal net, or lattice. This lattice in diffraction space is called the reciprocal lattice. 

Considerable interest was aroused by the finding of Wieme and Lauryssens (W16) in 1962 that there is a change in the electrophoretic isoenzyme pattern of lactate dehydrogenase in diseased human muscle. The major isoenzyme of lactate dehydrogenase in most normal muscles moves slowest on electrophoresis (LDH 5), but in myopathic muscle the proportion of LDH 5 may be considerably reduced. This finding has been confirmed and extended by numerous workers, utilizing various techniques for isoenzyme differentiation (e.g., BIO, E5). The abnormal pattern is seen in most, but not all, cases of Duchenne dystrophy and in a variety of other muscular disorders. It may be evident in the very early stages of Duchenne dystrophy (P2) and is seen even in some female carriers of the disease (E3). 

E3. Emery, A. E. H., Electrophoretic pattern of lactate dehydrogenase in carriers and patients with Duehenne muscular dystrophy. Nature (London) 201,1044-1045 (1964). 

The diagnosis of organ disease is aided by measurement of a number of enzymes characteristic of that tissue or organ. Most tissues have characteristic enzyme patterns (Table 8-2) that may be reflected in the relative serum concentrations of the respective enzymes in disease. The diseased tissue can be further identified by determination of the isoenzyme pattern of one of these enzymes (e.g., lactate dehydrogenase, creatine kinase) in the serum, since many tissues have characteristic isoenzyme distribution patterns for a given enzyme. For example, creatine kinase (CK) is a dimer composed of two subunits, M (for muscle) and B (for brain), that occur in three isoenzyme forms, BB(CKi), MB(CK2) and MM(CK3), which catalyze the reversible phosphorylation of creatine with adenosine triphosphate (ATP) as the phosphate donor ... 

Fig. 17. Typical patterns of Mn-SOD release into serum for two male patients with acute myocardial infarction. (A) A case with an unsuccessful reperfusion (B) successful case. The mean level for normal male adults is almost 100 ng/ml. Other enzymes AST, L-aspartate 2-oxoglutarate ALT, L-alanine 2-oxoglutarate CK, creatine kinase LDH, lactate dehydrogenase.

M31. McKenzie, D., and Henderson, A. R., An artifact in lactate dehydrogenase isoenzyme patterns, assayed by fluorescence, occurring in the serum of patients with end-stage renal disease requiring maintenance hemodialysis. Clin. Ckim. Acta 70, 33-336 (1976). 

A profile of the serum isoenzymes of lactate dehydrogenase, (a) The pattern of LDH isoenzymes from a normal individual, (b) The pattern of LDH isoenzymes of an individual suffering from a myocardial infarction. 

Lactate produced by smooth muscle of swine carotid artery has been shown to be quantitatively derived from extracellular glucose, whereas gly-cogenolysis provides carbohydrates for oxidative metabolism (Lynch and Paul, 1983, 1987). Interestingly, the lactate dehydrogenase (LDH) isoenzyme pattern varies between slow (aorta) and fast (portal vein and bladder) smooth muscle in rat, such that aorta has a higher proportion of the H subunit, which has a higher affinity for lactate and is product-inhibited at lower concentrations of lactate (Malmqvistef flZ., 1991). Thus the slow aortic muscle is more adapted for oxidative metabolism than the faster bladder and portal vein muscles. 

Fortunately, as the possible mechanisms proliferate, the analytical power of enzyme kinetics increases, since the kinetic patterns lead to diagnostic tests for various mechanisms. The task of unravelling 2- and 3-substrate mechanisms began in earnest in the 1950 s with the theoretical framework laid by Alberty [45,46], Dalziel [47,48] and Cleland [49-51]. The detailed experimental investigations of horse liver alcohol dehydrogenase by Theorell, Dalziel and their collaborators [42,52] and of beef heart lactate dehydrogenase by Schwert s group [53] were important milestones. 

Schultz, A. E., K. P. Gungaga, J. G. Wagner, C. M. Hoorn, W. R. Moorhead, and R. A. Roth. 1994. Lactate dehydrogenase activity and isozyme patterns in tissues and bronchoalveo-lar lavage fluid from rats treated with monocrotaline pyrrole. Toxicology and Applied... 

Fig. 7.8. Ribbon drawing showing the arrangement of secondary structures into a three-dimensional pattern in domain 1 of lactate dehydrogenase. The individual polyjjeptide strands in the six-stranded P-sheet are shown with arrows. Different strands are connected by helices and by nonrepetitive structures (mrns, coils and loops), shown in blue. This domain is the nucleotide binding fold. NAD is bound to a site created by the helices (upper left of figure.) (Modified from Richardson JS. Adv Protein Chem. The anatomy and taxonomy of protein structure 1981 34 167).

Serum levels of lactate dehydrogenase (LDH) were formerly used to diagnose an acute Ml. LDH is present in cells as a tetramer of four identical, or nearly identical, subunits. Each subunit is a separate polypeptide chain with a molecular weight of 35 kD (approximately 35,000 g/mole). These subunits are present as two isoforms, the H isoform (for heart) and the M isoform (for skeletal muscle). Although the heart produces principally the H4 form (four H subunits combined into a tetramer) and skeletal muscles produce principally the M4 isoform, the heart, skeletal muscle, and other tissues produce several intermediate combinations (e.g., H3M, HjMj). These tetrameric isoforms all have similar activities, but the individual monomeric subunits are inactive. Measurements of LDH isozymes in the serum are no longer used for diagnosis of a recent Ml because the enzyme is large, released slowly, and the isozyme pattern is not as specific for the heart as is CK. 

Gupta, R. C.. Goad. J. T., and Kadel. W. L. (1991b). In vivo alterations in lactate dehydrogenase (LDH) and LDH isoenzymes patterns by acute carbofuran intoxication, Arch. miiwi, Coniam, Toxicol. 21, 263-269,... 

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