Malate dehydrogenase in phototrophic purple bacteria: purification, molecular weight, and quaternary structure

Tayeh, Mahmoud A.; Madigan, Michael T.

doi:10.1128/jb.169.9.4196-4202.1987

Cited by 57 publications

(45 citation statements)

References 31 publications

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“…N-terminal amino acid sequence and composition. Figure 2 shows the sequences of residues in the N terminus of MDH from C. vibrioforme (40 positions (31). One disadvantage of the method, however, is the relatively high concentration of NAD required to elute MDH from the ligand.…”

Section: Methodsmentioning

confidence: 99%

“…Dimeric and tetrameric forms are represented in both mesophilic and thermophilic bacteria (20,23,28,31,39) and within the same genus (30). The (34).…”

Section: Methodsmentioning

confidence: 99%

“…The molecular weight (MW) of small MDHs is in the range from 60,000 to 74,000, and that of the large form is between 117,000 and 146,000. All MDHs studied so far are homogeneous in subunit composition, with a subunit MW in the range from 30,000 to 38,000 (1,16,17,23,30,31). The small enzymes are thus dimers, and the large enzymes are thus tetramers.…”

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confidence: 99%

“…The small enzymes are thus dimers, and the large enzymes are thus tetramers. The large form is widespread among the purple nonsulfur bacteria, though the small form is also represented (31). The MDH of Chloroflexus aurantiacus is a tetramer (23).…”

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confidence: 99%

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Malate dehydrogenase from Chlorobium vibrioforme, Chlorobium tepidum, and Heliobacterium gestii: purification, characterization, and investigation of dinucleotide binding by dehydrogenases by use of empirical methods of protein sequence analysis

1992

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Malate dehydrogenase (MDH; EC 1.1.1.37) from strain NCIB 8327 of the green sulfur bacterium Chlorobium vibrioforme was purified to homogeneity by triazine dye affinity chromatography followed by gel filtration. Purification of MDH gave an approximately 1,000-fold increase in specific activity and recoveries of typically 15 to 20%. The criteria of purity were single bands on sodium dodecyl sulfate (SDS) and nondenaturing polyacrylamide electrophoresis (PAGE) and the detection of a single N terminus in an Edman degradation analysis. MDH activity was detected in purified preparations by activity staining of gels in the direction of malate oxidation. PAGE and gel filtration (Sephadex G-100) analyses showed the native enzyme to be a dimer composed of identical subunits both at room temperature and at 4 degrees C. The molecular weight of the native enzyme as estimated by gel filtration was 77,000 and by gradient PAGE was 74,000. The subunit molecular weight as estimated by SDS-gradient PAGE was 37,500. N-terminal sequences of MDHs from C. vibrioforme, Chlorobium tepidum, and Heliobacterium gestii are presented. There are obvious key sequence similarities in MDHs from the phototrophic green bacteria. The sequences presented probably possess a stretch of amino acids involved in dinucleotide binding which is similar to that of Chloroflexus aurantiacus MDH and other classes of dehydrogenase enzymes but unique among MDHs.

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Section: Methodsmentioning

confidence: 99%

“…Dimeric and tetrameric forms are represented in both mesophilic and thermophilic bacteria (20,23,28,31,39) and within the same genus (30). The (34).…”

Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

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confidence: 99%

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Malate dehydrogenase from Chlorobium vibrioforme, Chlorobium tepidum, and Heliobacterium gestii: purification, characterization, and investigation of dinucleotide binding by dehydrogenases by use of empirical methods of protein sequence analysis

1992

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“…The three genes cvmdh, ctmdh, and cv71tmdh were expressed in E. coli BL21(DE3) by induction with 0.4 mM isopropyl-␤-D-thiogalacto-pyranoside (IPTG) (37), when the optical density at 600 nm of the cultures was 0.6 to 0.7. After 180 min of induction, the cells were collected and the foreign MDH was purified as described previously (39). The fractions containing the highest activity were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), pooled, and used for further analysis.…”

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confidence: 99%

Malate dehydrogenase from the mesophile Chlorobium vibrioforme and from the mild thermophile Chlorobium tepidum: molecular cloning, construction of a hybrid, and expression in Escherichia coli

1996

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The genes (mdh) encoding malate dehydrogenase (MDH) from the mesophile Chlorobium vibrioforme and the moderate thermophile C. tepidum were cloned and sequenced, and the complete amino acid sequences were deduced. When the region upstream of mdh was analyzed, a sequence with high homology to an operon encoding ribosomal proteins from Escherichia coli was found. Each mdh gene consists of a 930-bp open reading frame and encodes 310 amino acid residues, corresponding to a subunit weight of 33,200 Da for the dimeric enzyme. The amino acid sequence identity of the two MDHs is 86%. Homology searches using the primary structures of the two MDHs revealed significant sequence similarity to lactate dehydrogenases. A hybrid mdh was constructed from the 3 part of mdh from C. tepidum and the 5 part of mdh from C. vibrioforme. The thermostabilities of the hybrid enzyme and of MDH from C. vibrioforme and C. tepidum were compared.Malate dehydrogenase (MDH) catalyzes the reversible conversion of malate to oxaloacetate, using NAD as cofactor. In most organisms, MDH is an essential enzyme in carbon metabolism, and in green sulfur bacteria, it is part of the reductive tricarboxylic acid cycle (9).Complete amino acid sequence data for MDH are available from many species within the Bacteria, Archaea, and Eucarya. Based on sequence similarity, Goward and Nicholls (15) have suggested that MDH has diverged into two distinct phylogenetic groups, one group that includes cytoplasmic MDH, chloroplast MDH, and MDH from Thermus flavus, and a second group that includes MDHs with similarity to lactate dehydrogenase (LDH). In the latter group, the mitochondrial MDH and eubacterial MDH are in one cluster and the more LDHlike (archaeal) Haloarcula marismortui MDH is in another.Although the primary structures of the various MDHs and LDHs are different, their tertiary structures are similar. The dehydrogenases have two domains; the structurally conserved N-terminal dinucleotide-binding domain, and the C-terminal catalytic domain, which differs among the different dehydrogenases. The dinucleotide-binding domain of the LDHs and MDHs that use NAD as a cofactor contains a characteristic glycine motif. In LDH, this is GXGXXG, whereas for most known MDHs, it is GAXGXX(G/A). However, MDHs from all photosynthetic bacteria studied so far have the glycine motif typical of LDH (4, 29).The amino acid residues important for substrate and coenzyme binding, catalysis, and subunit assembly have been identified in several MDHs and LDHs (3,27,36,42). With regard to thermostability, LDH from Bacillus species has been extensively studied (21,41,45,46), and studies have also been performed with MDH from Escherichia coli (14) and from the thermophilic Thermus aquaticus (6).In the present work, MDH and its corresponding gene from the two green sulfur bacteria Chlorobium tepidum and Chlorobium vibrioforme have been isolated, characterized, and compared. The green sulfur bacteria comprise a distinct phylogenetic group of photosynthetic species, which are related to other gro...

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Fungicidal activity of novel quinazolin‐6‐ylcarboxylates and mode of action on Botrytis cinerea

Yan

Chen

et al. 2023

Pest Management Science

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BACKGROUND Fungal diseases remain important causes of crop failure and economic losses. As the resistance toward current selective fungicides becomes increasingly problematic, it is necessary to develop efficient fungicides with novel chemotypes. RESULTS A series of novel quinazolin‐6‐ylcarboxylates which combined the structures of pyridine or heterocyclic motif and the N‐(3‐chloro‐4‐fluorophenyl)quinazolin‐4‐amine moiety, a binding group of ATP‐binding site of gefitinib, were evaluated for their fungicidal activity on different phytopathogenic fungi. Most of these compounds showed excellent fungicidal activities against Botrytis cinerea and Exserohilum rostratum, especially compound F17 displayed the highest activity with EC50 values as 3.79 μg mL−1 against B. cinerea and 2.90 μg mL−1 against E. rostratum, which was similar to or even better than those of the commercial fungicides, such as pyraclostrobin (EC50, 3.68, 17.38 μg mL−1) and hymexazol (EC50, 4.56, 2.13 μg mL−1). Moreover, compound F17 significantly arrested the lesion expansion of B. cinerea infection on tomato detached leaves and strongly suppressed grey mold disease on tomato seedlings in greenhouse. The abilities of compound F17 to induce cell apoptosis of the non‐germinated spores, to limit oxalic acid production, to reduce malate dehydrogenase (MDH) expression, and to block the active pocket of MDH protein were demonstrated in B. cinerea. CONCLUSION The novel quinazolin‐6‐ylcarboxylates containing ATP‐binding site‐directed moiety, especially compound F17, could be developed as a potential fungicidal candidate for further study. © 2023 Society of Chemical Industry.

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Malate dehydrogenase in phototrophic purple bacteria: purification, molecular weight, and quaternary structure

Cited by 57 publications

References 31 publications

Malate dehydrogenase from Chlorobium vibrioforme, Chlorobium tepidum, and Heliobacterium gestii: purification, characterization, and investigation of dinucleotide binding by dehydrogenases by use of empirical methods of protein sequence analysis

Malate dehydrogenase from Chlorobium vibrioforme, Chlorobium tepidum, and Heliobacterium gestii: purification, characterization, and investigation of dinucleotide binding by dehydrogenases by use of empirical methods of protein sequence analysis

Malate dehydrogenase from the mesophile Chlorobium vibrioforme and from the mild thermophile Chlorobium tepidum: molecular cloning, construction of a hybrid, and expression in Escherichia coli

Fungicidal activity of novel quinazolin‐6‐ylcarboxylates and mode of action on Botrytis cinerea

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