Enzymes lactate dehydrogenase

F28. Freier, E. F., Genetic and conformational heterogeneity of lactate dehydrogenase enzymes. Texas Bept. Biol. Med. 24, 375-385 (1966). 

Microbial contamination of milk can be tested through determination of l- and D-lactate. Two amperometric biosensors based on the l- and D-lactate dehydrogenase enzymes have been proposed for l- and D-lactate assay.119 They assure a continuous monitorizing of l- and D-lactate in milk, and they also assure good reliability of the analytical information. 

Other examples of amperometric enzyme electrodes based on the measurement of oxygen or hydrogen peroxide include electrodes for the measurement of galactose in blood (galactose oxidase,enzyme), oxalate in urine (oxalate oxidase), and cholesterol in blood serum (cholesterol oxidase). Ethanol is determined by reacting with a cofactor, nicotinamide adenine dinucleotide (NAD" ) in the presence of the enzyme alcohol dehydrogenase to produce the reduced form of NAD", NADH, which is electrochemically oxidized. Lactate in blood is similarly determined (lactate dehydrogenase enzyme). 

The genes for both of the lactate dehydrogenase enzymes, l-LDH and d-LDH, that control lactic acid stereospecificity in the lactobacilli have been cloned and sequenced. The l-LDH enzyme from a variety of lactic acid bacteria forms a closely related class entirely distinct from the d-LDH enzymes which form a second closely related group of proteins. With broad host-range integration vectors now available, it is possible to... 

Enzyme catalyzed reductions of carbonyl groups are more often than not com pletely stereoselective Pyruvic acid for example is converted exclusively to (5) (+) lactic acid by the lactate dehydrogenase NADH system (Section 15 11) The enantiomer... 

Multienzyme electrodes can increase sensitivity from micromolar to nanomolar detection levels (53,57). In this case the substrate is converted to a detectable product by one enzyme, then that product is recycled into the initial substrate by another enzyme resulting in an amplification of the response signal. For example, using lactate oxidase and lactate dehydrogenase immobilized in poly(vinyl chloride), an amplification of 250 was obtained for the detection oflactate (61). 

Jervis used porous silica coated with chemisorbed polyacrylhydrazide for immobilization of adenosine monophosphate (AMP) [117]. After periodate oxidation of its ribose residue the ligand was coupled to the carrier and used for isolation of lactate dehydrogenase from rabbit muscle. The specific capacity was 2 mg of protein/g adsorbent with a ligand content of 10 pmol/g, whereas recovery of enzymatic activity after elution was 85%. Hipwell et al. [118] found that for effective binding of lactate dehydrogenases on AMP-o-aminoalkyl-Sepharose the spacer arm length required at least 4 methylene links. Apparently, a macromolecule of polyacrylhydrazide acts itself like an extended spacer arm and thus allow AMP to bind the enzyme. 

Directed evolution of enzymes has been used to improve the reducing function of the enzymes. For example, this method was used to eliminate the cofactor requirement of B. stearothermophillus lactate dehydrogenase, which is activated in the presence of fructose 1,6-bisphosphate [12]. The activator is expensive and representative of the sort of cofactor complications that are undesirable in industrial processes. Three rounds of random mutagenesis and screening produced a mutant that is almost fully... 

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). 

L-Lactate dehydrogenase is a tetrameric enzyme whose four subunits occur in two isoforms, designated H (for... 

Assay of plasma enzymes aids diagnosis and prognosis. For example, a myocardial infarction elevates serum levels of lactate dehydrogenase isozyme I,. 

CK catalyzes the reversible phosphorylation of creatine in the presence of ATP and magnesium. When creatine phosphate is the substrate, the resulting creatine can be measured as the ninhydrin fluorescent compound, as in the continuous flow Auto Analyzer method. Kinetic methods based on coupled enzymatic reactions are also popular. Tanzer and Gilvarg (40) developed a kinetic method using the two exogenous enzymes pyruvate kinase and lactate dehydrogenase to measure the CK rate by following the oxidation of NADH. In this procedure the main reaction is run in a less favorable direction. 

Studies of release of noradrenaline from sympathetic neurons provided the first convincing evidence that impulse (Ca +)-dependent release of any transmitter depended on vesicular exocytosis. Landmark studies carried out in the 1960s, using the perfused cat spleen preparation, showed that stimulation of the splenic nerve not only led to the detection of noradrenaline in the effluent perfusate but the vesicular enzyme, DpH, was also present. As mentioned above, this enzyme is found only within the noradrenaline storage vesicles and so its appearance along with noradrenaline indicated that both these factors were released from the vesicles. By contrast, there was no sign in the perfusate of any lactate dehydrogenase, an enzyme that is found only in the cell cytosol. The processes by which neuronal excitation increases transmitter release were described in Chapter 4. 

In addition to enzyme activity, the concentration of an nonelectroactive substrate can be determined electrochemically by this technique. By keeping the substrate (analyte) the limiting reagent, the amount of product produced is directly related to the initial concentration of substrate. Either kinetic or equilibrium measurements can be used. Typically an enzyme which produces NADH is used because NADH is readily detected electrochemically. Lactate has been detected using lactate dehydrogenase, and ethanol and methanol detected using alcohol dehydrogenase... 

Many dehydrogenase enzymes catalyze oxidation/reduction reactions with the aid of nicotinamide cofactors. The electrochemical oxidation of nicotinamide adeniiw dinucleotide, NADH, has been studied in depthThe direct oxidation of NADH has been used to determine concentration of ethanol i s-isv, i62) lactate 157,160,162,163) pyTuvate 1 ), glucose-6-phosphate lactate dehydrogenase 159,161) alanine The direct oxidation often entails such complications as electrode surface pretreatment, interferences due to electrode operation at very positive potentials, and electrode fouling due to adsorption. Subsequent reaction of the NADH with peroxidase allows quantitation via the well established Clark electrode. 

Deficiencies of enzymes involved in glycolysis, the hexose monophosphate pathway, the closely related glutathione metabolism and synthesis, and nucleotide metabolism have emerged as causes of hereditary nonspherocytic hemolytic anemias (Table 1) (F10, Fll, M27). Some enzyme deficiencies, such as diphospho-glycerate mutase deficiency, lactate dehydrogenase deficiency, and NADH cy-... 

---