scholarly journals Structure and Function of Benzylsuccinate Synthase and Related Fumarate-Adding Glycyl Radical Enzymes

2016 ◽  
Vol 26 (1-3) ◽  
pp. 29-44 ◽  
Author(s):  
Johann Heider ◽  
Maciej Szaleniec ◽  
Berta M. Martins ◽  
Deniz Seyhan ◽  
Wolfgang Buckel ◽  
...  
Keyword(s):  
Anaerobic Metabolism ◽  
Biochemical Properties ◽  
Initial Structure ◽  
Toluene Degradation ◽  
Methyl Group ◽  
Glycyl Radical ◽  
Benzylsuccinate Synthase ◽  
And Function ◽  
Glycyl Radical Enzyme ◽  
Radical Type

The pathway of anaerobic toluene degradation is initiated by a remarkable radical-type enantiospecific addition of the chemically inert methyl group to the double bond of a fumarate cosubstrate to yield <i>(R)</i>-benzylsuccinate as the first intermediate, as catalyzed by the glycyl radical enzyme benzylsuccinate synthase. In recent years, it has become clear that benzylsuccinate synthase is the prototype enzyme of a much larger family of fumarate-adding enzymes, which play important roles in the anaerobic metabolism of further aromatic and even aliphatic hydrocarbons. We present an overview on the biochemical properties of benzylsuccinate synthase, as well as its recently solved structure, and present the results of an initial structure-based modeling study on the reaction mechanism. Moreover, we compare the structure of benzylsuccinate synthase with those predicted for different clades of fumarate-adding enzymes, in particular the paralogous enzymes converting <i>p</i>-cresol, 2-methylnaphthalene or <i>n</i>-alkanes.

scholarly journals Stable Isotope Fractionation Caused by Glycyl Radical Enzymes during Bacterial Degradation of Aromatic Compounds

2004 ◽  
Vol 70 (5) ◽  
pp. 2935-2940 ◽  
Author(s):  
Barbara Morasch ◽  
Hans H. Richnow ◽  
Andrea Vieth ◽  
Bernhard Schink ◽  
Rainer U. Meckenstock
Keyword(s):  
Stable Isotope ◽  
Carbon Isotope ◽  
Isotope Fractionation ◽  
Toluene Degradation ◽  
Fractionation Factor ◽  
Glycyl Radical ◽  
Stable Isotope Fractionation ◽  
Benzylsuccinate Synthase ◽  
Glycyl Radical Enzyme

ABSTRACT Stable isotope fractionation was studied during the degradation of m-xylene, o-xylene, m-cresol, and p-cresol with two pure cultures of sulfate-reducing bacteria. Degradation of all four compounds is initiated by a fumarate addition reaction by a glycyl radical enzyme, analogous to the well-studied benzylsuccinate synthase reaction in toluene degradation. The extent of stable carbon isotope fractionation caused by these radical-type reactions was between enrichment factors (ε) of −1.5 and −3.9, which is in the same order of magnitude as data provided before for anaerobic toluene degradation. Based on our results, an analysis of isotope fractionation should be applicable for the evaluation of in situ bioremediation of all contaminants degraded by glycyl radical enzyme mechanisms that are smaller than 14 carbon atoms. In order to compare carbon isotope fractionations upon the degradation of various substrates whose numbers of carbon atoms differ, intrinsic ε (εintrinsic) were calculated. A comparison of εintrinsic at the single carbon atoms of the molecule where the benzylsuccinate synthase reaction took place with compound-specific ε elucidated that both varied on average to the same extent. Despite variations during the degradation of different substrates, the range of ε found for glycyl radical reactions was reasonably narrow to propose that rough estimates of biodegradation in situ might be given by using an average ε if no fractionation factor is available for single compounds.

scholarly journals Benzylsuccinate Synthase is Post-Transcriptionally Regulated in the Toluene-Degrading Denitrifier Magnetospirillum sp. Strain 15-1

2020 ◽  
Vol 8 (5) ◽  
pp. 681 ◽  
Author(s):  
Ingrid Meyer-Cifuentes ◽  
Sylvie Gruhl ◽  
Sven-Bastiaan Haange ◽  
Vanessa Lünsmann ◽  
Nico Jehmlich ◽  
...  
Keyword(s):  
Transcriptional Control ◽  
Toluene Degradation ◽  
Regulation Of Expression ◽  
Glycyl Radical ◽  
Copy Numbers ◽  
Catalytically Active ◽  
Oxygen Sensitive ◽  
Benzylsuccinate Synthase ◽  
Glycyl Radical Enzyme ◽  
In Cells

The facultative denitrifying alphaproteobacterium Magnetospirillum sp. strain 15-1 had been isolated from the hypoxic rhizosphere of a constructed wetland model fed with toluene. This bacterium can catabolize toluene anaerobically but not aerobically. Here, we used strain 15-1 to investigate regulation of expression of the highly oxygen-sensitive glycyl radical enzyme benzylsuccinate synthase, which catalyzes the first step in anaerobic toluene degradation. In cells growing aerobically with benzoate, the addition of toluene resulted in a ~20-fold increased transcription of bssA, encoding for the catalytically active subunit of the enzyme. Under anoxic conditions, bssA mRNA copy numbers were up to 129-fold higher in cells growing with toluene as compared to cells growing with benzoate. Proteomics showed that abundance of benzylsuccinate synthase increased in cells growing anaerobically with toluene. In contrast, peptides of this enzyme were never detected in oxic conditions. These findings show that synthesis of benzylsuccinate synthase was under stringent post-transcriptional control in the presence of oxygen, which is a novel level of regulation for glycyl radical enzymes.

scholarly journals Determinants for Substrate Recognition in the Glycyl Radical Enzyme Benzylsuccinate Synthase Revealed by Targeted Mutagenesis

ACS Catalysis ◽  
2021 ◽  
pp. 3361-3370
Author(s):  
Iryna Salii ◽  
Maciej Szaleniec ◽  
Ammar Alhaj Zein ◽  
Deniz Seyhan ◽  
Anna Sekuła ◽  
...  
Keyword(s):  
Substrate Recognition ◽  
Targeted Mutagenesis ◽  
Glycyl Radical ◽  
Benzylsuccinate Synthase ◽  
Glycyl Radical Enzyme

Identification of FeS clusters in the glycyl-radical enzyme benzylsuccinate synthase via EPR and Mössbauer spectroscopy

2011 ◽  
Vol 17 (1) ◽  
pp. 49-56 ◽  
Author(s):  
Markus Hilberg ◽  
Antonio J. Pierik ◽  
Eckhard Bill ◽  
Thorsten Friedrich ◽  
Marie-Luise Lippert ◽  
...  
Keyword(s):  
Mössbauer Spectroscopy ◽  
Mossbauer Spectroscopy ◽  
Mößbauer Spectroscopy ◽  
Glycyl Radical ◽  
Benzylsuccinate Synthase ◽  
Glycyl Radical Enzyme

scholarly journals Anaerobic Toluene Catabolism of Thauera aromatica: the bbs Operon Codes for Enzymes of β Oxidation of the Intermediate Benzylsuccinate

2000 ◽  
Vol 182 (2) ◽  
pp. 272-277 ◽  
Author(s):  
Birgitta Leuthner ◽  
Johann Heider
Keyword(s):  
Initial Reaction ◽  
Transcriptional Level ◽  
Beta Oxidation ◽  
Oxidation Pathway ◽  
Glycyl Radical ◽  
Thauera Aromatica ◽  
Coa Transferase ◽  
Benzylsuccinate Synthase ◽  
Glycyl Radical Enzyme ◽  
Induced Proteins

ABSTRACT The pathway of anaerobic toluene oxidation to benzoyl coenzyme A (benzoyl-CoA) consists of an initial reaction catalyzed by benzylsuccinate synthase, a glycyl radical enzyme adding the methyl group of toluene to the double bond of a fumarate cosubstrate, and a subsequent β-oxidation pathway of benzylsuccinate. Benzylsuccinate synthase has been studied in some detail, whereas the enzymes participating in β oxidation of benzylsuccinate are unknown. We have investigated these enzymes by analyzing substrate-induced proteins in toluene-grown cells. Toluene-induced proteins were identified and N-terminally sequenced. Nine of these proteins are encoded by an 8.5-kb operon consisting ofbbs (beta-oxidation of benzylsuccinate) genes whose products are apparently involved in the β-oxidation pathway of benzylsuccinate. Two of the genes, bbsE andbbsF, code for the subunits of a succinyl-CoA:benzylsuccinate CoA-transferase whose activity was previously detected in toluene-grown Thauera aromatica. The bbsG gene codes for a specific benzylsuccinyl-CoA dehydrogenase, as confirmed by overexpression of the gene in Escherichia coli and detection of enzyme activity. The further enzymes of the pathway are probably encoded bybbsH (enoyl-CoA hydratase), bbsCD(3-hydroxyacyl-CoA dehydrogenase), and bbsB (3-oxoacyl-CoA thiolase). The operon contains two additional genes, bbsAand bbsI, for which no obvious function could be derived. The bbs operon is expressed only in toluene-grown cells and is regulated at the transcriptional level. Promoter mapping revealed a transcription start site upstream of the bbsA gene. This represents the first known promoter site in Thauera spp.

Identification of a large complex containing the integrin alpha 6 beta 1 laminin receptor in neural retinal cells

1994 ◽  
Vol 107 (11) ◽  
pp. 3165-3172 ◽  
Author(s):  
I. de Curtis ◽  
G. Gatti
Keyword(s):  
Biochemical Properties ◽  
Cytoskeletal Proteins ◽  
Laminin Receptor ◽  
Retinal Neurons ◽  
Differential Distribution ◽  
Gradient Distribution ◽  
Large Complex ◽  
Beta 1 Integrin ◽  
Integrin Alpha 6 ◽  
And Function

Integrin alpha 6 beta 1 is a laminin receptor involved in adhesion and neurite extension of retinal neurons on laminin. The present study was carried out to identify potential interactions between the alpha 6 beta 1 receptor and cellular proteins that may be involved in integrin signaling and function. For this purpose we have used a biochemical approach based on the solubilization of retinal neurons cultured on laminin with nonionic detergents, followed by centrifugation on sucrose velocity gradients. Analysis of the distribution of the alpha 6 and beta 1 integrin subunits in the gradients showed that they migrate as a large complex after extraction of cells with octylglucoside, but not after Triton X-100 extraction. Cytoskeletal proteins known to localize in adhesion plaques did not comigrate with alpha 6 beta 1 in octylglucoside gradients, while a set of polypeptides whose tyrosine phosphorylation was enhanced by culture on laminin colocalized with alpha 6 beta 1 on the gradients after octylglucoside solubilization. Culture of retinal neurons on bovine serum albumin, a nonadhesive substratum, partially affected the gradient distribution of the receptor after octylglucoside extraction. Furthermore, analysis of the gradient distribution of two alternatively spliced isoforms of the alpha 6 subunit, alpha 6-cytoA and alpha 6-cytoB, showed that the effect of non-adhesion on the sedimentation properties of the two integrin alpha 6 isoforms was more dramatic for alpha 6-cytoB than alpha 6-cytoA. These differences in the sedimentation behaviour indicate distinct biochemical properties of the two alpha 6 isoforms that, together with previous observations on their differential distribution in the developing retina, may reflect functional specificities.

Sign in / Sign up

Export Citation Format

Share Document