Mycoplasma laidlawii possesses a single glutamate dehydrogenase (GDH) with dual coenzyme specificity [specificity for nicotinamide adenine dinucleotide (H) and nicotinamide adenine dinucleotide phosphate (H) ]. A purification procedure is reported which results in an enzyme preparation with a specific activity of 79.5 units/mg and which displays only one significant protein band after gel electrophoresis. This one band was determined, by activity staining, to have all of the GDH nucleotide specificities. The molecular weight of the enzyme is 250,000 + 10%, and it has a subunit size of about 48,000. The enzyme exhibits measurable activity with aspartate and pyruvate but is inactive with eight other possible substrates. Purine nucleotides do not affect the activity. The Km for reduced nicotinamide adenine dinucleotide was 1.8 x 10-4 M. The optimal substrate concentrations and pH optimum for each of the respective GDH activities are also reported. MATERIALS AND METHODS Chemicals. a-Ketoglutaric acid, NAD, NADPH, ribonuclease (all A grade) and deoxyribonuclease (B grade) were obtained from Calbiochem. Phenol reagent (Folin-Ciocalteau) was from Fisher Scientific Co. Chemicals for acrylamide gel electrophoresis were all products of Matheson, Coleman, and Bell. Phenazine methosulfate, MTT tetrazolium [3(4,5 dimethyl thiazolyl-2)-2, 5-diphenyl tetrazolium bromide], tris(hydroxymethyl) aminomethane (Tris), NADP (Sigma grade), and NADH (grade 111) were obtained from Sigma Chemical Co. Sepharose 6B and diethylaminoethyl (DEAE) Sephadex A-50 were obtained from Pharmacia Fine Chemicals, Inc. Ammonium sulfate, A.R. was from Mallinckrodt Chemical Works. DEAE cellulose DE.11 was obtained from Reeve Angel, Inc. Crude extract preparation. M. laidlawii, strain A, culture and osmotic lysis procedure were as described previously (4). The organisms were extracted once with 10-3 M ethylenediaminetetraacetic acid (EDTA), pH 7.5, followed by two more extractions with demineralized water. The extracts were then combined and subjected to the enzyme fractionation procedure described later.

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Prostatic acid phosphatase and alkaline phosphatase values in bone marrow were correlated with skeletal surveys and diagnoses during a six-month study. In cases of biopsy-proven adenocarcinoma of the prostate, bone marrow prostatic acid phosphatase was the most consistently abnormal value. Diagnoses other than prostatic cancer involving the bone marrow, e.g., myeloma and leukemias, were associated with elevated prostatic acid phosphatase and alkaline phosphatase values. In cases in which the bone marrow was not involved by metastasis, these values were normal. Bone marrow prostatic acid phosphatase assay was found to be a very good tool for detecting early osseous metastases from any site, including prostatic adenocarcinoma.

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