Localization of Reduced Nicotinamide Adenine Dinucleotide Oxidase Activity in Acholeplasma and Mycoplasma Species

Pollack, Jonathan D.

doi:10.1099/00207713-25-2-108

Cited by 27 publications

(15 citation statements)

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“…Cells were harvested in midlog growth (18 to 72 h), washed, hypotonically lysed, and centrifugally fractionated, as described previously for enzyme location studies (13,14). Acholeplasmal extracts were not subjected to further disruptive procedures, but all spiroplasmal and mycoplasmal preparations were also exposed to 65 W of sonic oscillation with a model 350 Branson Sonifier (Heat Systems Co., New York, N.Y.) for three 5-s bursts while they were in a wet ice bath.…”

Section: Methodsmentioning

confidence: 99%

Pyrimidine Deoxyribonucleotide Metabolism in Members of the Class Mollicutes

Williams

Pollack²

1985

International Journal of Systematic Bacteriology

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Cell extracts from six Acholeplasma species, six Mycoplasma species, and Spiroplasma Jlwicola 23-6T (T = type strain) were examined for enzyme activities of pyrimidine deoxyribonucleotide metabolism. A11 of these organisms had thymidine kinase and thymidine phosphorylase activities, and all lacked deoxycytidine triphosphatase activity. The 13 members of the Mollicutes were separated into three groups by the presence or absence of the following four enzyme activities: (i) the adenosine triphosphate-insensitive deoxyuridine triphosphate-specific hydrolyzing deoxyuridine triphosphatase, (ii) a deoxyuridine monophosphate phosphatase, (iii) deoxycytidine deaminase, and (iv) deoxycytidine monophosphate deaminase. Five of the six Acholeplasma species (all Acholeplasma species except Acholeplasma florum LIT) had all four enzymatic activities. The six Mycoplasma species only had the deoxycytidine and deoxycytidine monophosphate deaminase activities. The only two plant isolates studied, A . Jlorum LIT and S . Jloricola 23-6T, lacked all four enzymatic activities.The class Mollicutes contains two orders. The order Mycoplasmatales is composed of two families, the Mycoplasmataceae, which has two genera (Mycoplasma and Ureaplasma), and the Spiroplasmataceae, which has o n e genus ( S p i r o p l a s m a ) (4). The second order, A c h o l e p l a s m a t a l e s , c o n t a i n s o n e f a m i l y , t h e Achdeplasmataceae, with one genus, Acholeplasma (5). A number of biochemical, nutritional, and morphological characteristics have been used to distinguish these genera and families. They include sterol requirement for growth, genome sizes, ability to hydrolyze urea, localization of reduced nicotinamide adenine dinucleotide oxidase, and presence of helical forms during growth (4, 5). However, no one has distinguished genera or families of the Mollicutes by the presence or absence of enzymes involved in pyrimidine deoxyribonucleotide metabolism.Recently, we characterized the adenosine triphosphate (ATP)-insensitive highly specific deoxyuridine triphosphate ( d U T P ) -h y d r 01 y zi ng d e o x y uri d i n e t r i p h o s p h a t e nucleotidohydrolase (dUTPase; EC 3.6.1.23) from Acholeplasma laidlawii B-PG9 (17). In the present study, we examined five other Acholeplasma species, six Mycoplasma species, and one Spiroplasma species for dUTPase and o t h e r e n z y m e a c t i v i t i e s involved in pyrimidine deoxyribonucleotide metabolism. Our data suggest that it may be possible, with one exception, to distinguish these genera based upon the presence or absence of the ATP-insensitive dUTP-specific dUTPase (17), deoxyuridine monophosphate (dUMP) phosphatase, deoxycytidine (dC) deaminase, and deoxycytidine monophosphate (dCMP) deaminase activities in cell extracts. (56.6 Ci/mmol) were purchased from Amersham Corp., Arlington Heights, Ill. Polyethyleneimine-cellulose thinlayer chromatography plates were purchased from Analtech, Newark, Del. MATERIALS AND METHODSOrganisms. Acholeplasma florum LIT (T = type strain) and Spiroplasm...

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Section: Methodsmentioning

confidence: 99%

Pyrimidine Deoxyribonucleotide Metabolism in Members of the Class Mollicutes

Williams

Pollack²

1985

International Journal of Systematic Bacteriology

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“…laidlawii B-PG9 (14,24). Purity of the fractions was monitored by localization of reduced nicotinamide adenine dinucleotide oxidase (13,14) and adenosine triphosphatase (EC 3.6.1.3) (24) activities. Assay.…”

Section: Methodsmentioning

confidence: 99%

Distinctions in Mollicutes Purine Metabolism: Pyrophosphate-Dependent Nucleoside Kinase and Dependence on Guanylate Salvage

Tryon

Pollack

1985

International Journal of Systematic Bacteriology

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Cell preparations of Acholeplasma jlorum LIT, Acholeplasma axanthum S743T, Acholeplasma granularum BTS-39T, Spiroplasma jloricola 23-6T, Mycoplasma gallisepticum S6, and Mycoplasma arginini G230T were examined for 13 cytoplasmic enzyme activities involved in the salvage and interconversion of nucleobases, nucleosides, and 5'-mononucleotides. The unique pyrophosphate-dependent nucleoside kinase activity known only in Acholeplasma laidlawii B-PG9 was found in A . axanthum and A . granularum. All the organisms could be divided into three groups based upon their patterns of purine salvage and interconversion activities. All the tested organisms, A. laidlawii, and apparently Mycoplasma mycoides subsp. mycoides lack the ability to synthesize guanylates from other purine mononucleotides, indicating that these members of the class Mollicutes can only salvage guanylates. The plant epiphytes A . florum and S . jloricola had an identical purine enzyme pattern which was different from those of all other members of the Mollicutes studied.Mollicutes (mycoplasmas) is a class of procaryotes characterized by lack of a cell wall, complex nutritional requirements, and small genome of generally low guanine-pluscytosine ratio. Nutritional studies on representatives from each of the three families within the class demonstrate the absolute requirement for nucleobases or nucleosides for growth (3, 5, 8, 11,[17][18][19][20][21]23). The nutritional requirement for purine precursors suggests the lack of a functional pathway for the de novo synthesis of purines and the dependence on salvage and interconversion enzymes for the biosynthesis of purine nucleotides (11, 24). Hamet et al. (6) examined purine salvage but not nucleotide interconversion activity in a number of Mycoplasma species. Only in Mycoplasma mycoides subsp. mycoides (11, 12), a species not generally available for study in the United States, and Acholeplasma laidlawii B-PG9 (24) have the pathways involved in purine salvage and interconversion been completely described.Here we describe our examination of the class Mollicutes for the unique pyrophosphate (PPJ-dependent nucleoside kinase activity which is only known in A. laidlawii B-PG9 (24). When first reported, we speculated that the PPidependent enzyme activity might prove useful in the study of the phylogenetic relationships of the Mollicutes. We now have studied six other representative Mollicutes for this PPi-dependent activity and for 12 other cytoplasmic enzyme activities involved in the salvage and interconversion of purine bases, ribonucleosides, and 5'-monophosphates. MATERIALS AND METHODSOrganisms. Acholeplasma florum LIT, Acholeplasma axanthum S743T, Acholeplasma granularum BTS-39T, and Spiroplasma joricola 23-6T were obtained from J. G. Tully, National Institutes of Health, Frederick, Md. Mycoplasma gallisepticum S6 and Mycoplasma arginini G230T were obtained from our stock collection.Media and growth conditions. All organisms were grown in ). Organisms were incubated statically at 37°C until mid-log phase of grow...

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“…Almost all the Mycoplasma species examined so far lack cytochromes but possess a flavin-terminated respiratory chain resembling that of lactobacilli (14,15). Moreover, in the sterol-requiring Mycoplasma species (9,10) and in Spiroplasma citri (S. Razin and S. Greenstein, unpublished data) the enzymatic system responsible for the transfer of electrons from reduced nicotinamide adenine dinucleotide (NADH) to oxygen is soluble and located in the cytoplasm. Only in the sterol-nonrequiring Acholeplasma species is this system membrane bound, as is generally the case in bacteria (9,10).…”

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“…The different location of the NADH oxidase in the Mycoplasma and Acholeplasma species, apart from its phylogenetic significance (9) ' Ptesent address: C.S.I.C. Centro de Investigaciones Biologicas, Instituto de Biologia Celular, Velazquez, 144, Madrid-6, Spain.…”

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Reduced nicotinamide adenine dinucleotide oxidase activity in membranes and cytoplasm of Acholeplasma laidlawii and Mycoplasma mycoides subsp. capri

Larraga¹,

Razin²

1976

J Bacteriol

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The properties of the membrane-bound reduced nicotinamide adenine dinucleotide (NADH) oxidase of Acholeplasma laidlawii were compared with those of the corresponding cytoplasmic activity of Mycoplasma mycoides subsp. capri. The striking differences in pH optima, susceptibility to inhibitors and detergents, and heat inactivation between the NADH oxidase activity, with oxygen as an electron acceptor, and the NADH oxidoreductase activity, with dichlorophenol indophenol (DCPIP) as an alternate electron acceptor, support the presence of more than one catalytic protein in both the membrane-bound and soluble enzyme systems. The detection of more than one band positive for the NADHnitroblue tetrazolium oxidoreductase reaction on electrophoresis of either the membranes of A. laidlawii or the cytoplasm of M. mycoides subsp. capri also points in the same direction. The membrane-bound enzyme system differed, however, from the soluble one because it had a lower ratio of oxidase activity to oxidoreductase activity, and because it was less susceptible to heat inactivation and more readily incorporated into reaggregated membranes. In addition, the specific activity of the membrane-bound enzyme system increased as the culture aged, whereas that of the soluble system decreased as the culture aged. It is suggested that the different location in the cell could be responsible for some of the differences between the membrane-bound NADH oxidase activity of A. laidlawii and that found in the cytoplasm of M. mycoides subsp. capri. the washed cells by osmotic lysis as described previ-827 on August 1, 2020 by guest

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Localization of Reduced Nicotinamide Adenine Dinucleotide Oxidase Activity in Acholeplasma and Mycoplasma Species

Cited by 27 publications

References 5 publications

Pyrimidine Deoxyribonucleotide Metabolism in Members of the Class Mollicutes

Pyrimidine Deoxyribonucleotide Metabolism in Members of the Class Mollicutes

Distinctions in Mollicutes Purine Metabolism: Pyrophosphate-Dependent Nucleoside Kinase and Dependence on Guanylate Salvage

Reduced nicotinamide adenine dinucleotide oxidase activity in membranes and cytoplasm of Acholeplasma laidlawii and Mycoplasma mycoides subsp. capri

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