ADH aus Rhodococcus erythropolis

Hummel W, Abokitse K, Gröger H. ADH aus Rhodococcus erythropolis. 2009

Towards a Large-Scale Asymmetric Reduction Process with Isolated Enzymes: Expression of an (S)-Alcohol Dehydrogenase in E. coli and Studies on the Synthetic Potential of this Biocatalyst

Hummel W, Abokitse K, Drauz K, Rollmann C, Gröger H. Towards a Large-Scale Asymmetric Reduction Process with Isolated Enzymes: Expression of an (S)-Alcohol Dehydrogenase in E. coli and Studies on the Synthetic Potential of this Biocatalyst. Adv. Synth. Catal. 2003;345:153-159

New feruloyl esterases to access phenolic acids from grass biomass

In the Sorangium cellulosum strain So ce56 genome, two putative esterase-encoding genes (loci sce1896 and sce8927) were cloned, expressed in Escherichia coli, and the resulting enzymes (designated ScFAE1 and ScFAE2) were used to assess the possible release of ferulic acid (FA) from triticale and wheat brans, and an aqueous fraction of steam-exploded wheat straw. The two polypeptides, sharing only 30% sequence identity, exhibit a typical catalytic Ser-Asp-His triad, a characteristic of \u3b1/\u3b2-hydrolase fold proteins. Both ScFAE1 (35 kDa) and ScFAE2 (34 kDa) were purified to apparent homogeneity and comparison of their kinetic parameters indicated an apparent higher affinity of ScFAE2 than ScFAE1 towards the various feruloyl substrates. This property was reflected by the observation that ScFAE2 was capable of yielding up to 85% of FA from destarched triticale bran. In the steam-exploded wheat sample, more than 85% yield of FA or p-coumaric acid was also effected by ScFAE2 without the decomposition of valuable chemical such as furfural. The two cloned FAEs represent the first of myxobacterial origin to be characterized and they are classified as new members of the type D family of FAEs.Peer reviewed: YesNRC publication: Ye

Structural analysis of Bacillus pumilus phenolic acid decarboxylase, a lipocalin-fold enzyme

Acta Crystallographica Section F: Structural Biology and Crystallization Communicationsis a rapid all-electronic journal, which provides a home for short communications onthe crystallization and structure of biological macromolecules. It includes four categoriesof publication: protein structure communications; nucleic acid structure communications;structural genomics communications; and crystallization communications. Structuresdetermined through structural genomics initiatives or from iterative studies suchas those used in the pharmaceutical industry are particularly welcomed. Section F isessential for all those interested in structural biology including molecular biologists, biochemists,crystallization specialists, structural biologists, biophysicists, pharmacologistsand other life scientists.Acta Crystallographica Section F: Structural Biology and Crystallization Communications est une revue enti\ue8rement \ue9lectronique \ue0 parution rapide renfermant de courtes communications sur la cristallisation et la structure de macromol\ue9cules biologiques. Elle se divise en quatre cat\ue9gories de communications, soit la structure des prot\ue9ines, la structure des acides nucl\ue9iques, la g\ue9nomique structurale et la cristallisation. Les structures d\ue9termin\ue9es par des initiatives en g\ue9nomique structurale ou \ue0 partir d\u2019\ue9tudes it\ue9ratives, comme celles employ\ue9es dans l\u2019industrie pharmaceutique, sont particuli\ue8rement bien accueillies. Cette revue constitue un incontournable pour tous ceux et celles qui s\u2019int\ue9ressent \ue0 la biologie structurale, soit les biologistes mol\ue9culaires, les biochimistes, les sp\ue9cialistes de la cristallisation, les biologistes structuraux, les biophysiciens, les pharmacologues et autres chercheurs en sciences de la vie.Peer reviewed: YesNRC publication: Ye

Thermostable feruloyl esterase for the bioproduction of ferulic acid from triticale bran

A putative \u3b1/\u3b2 hydrolase fold-encoding gene (locus tag TTE1809) from the genome of Thermoanaerobacter tengcongensis was cloned and expressed in Escherichia coli as a possible source of thermostable feruloyl esterase (FAE) for the production of antioxidant phenolic acids from biomass. Designated as TtFAE, the 33-kDa protein was purified to apparent homogeneity. The lipase-like sequence characteristics of TtFAE and its substrate specificity towards methyl ferulate, methyl sinapate, and methyl p-coumarate classify it as a new member of the type A FAEs. At 75\ub0C, the enzyme retained at least 95% of its original activity for over 80 min; at 80\ub0C, its half-life was found to be 50 min, rendering TtFAE a highly thermostable protein. Under different hydrolytic conditions, ferulic acid (FA) was shown to be released from feruloylated oligosaccharides prepared from triticale bran. An estimated recovery of 68 mg FA/100 g triticale bran was demonstrated by a 30% release of the total FA from triticale bran within a 5-h incubation period. Both the oxygen radical absorbing capacity values of the feruloylated oligosaccharides and free FA were also determined. Overall, this work introduces a new bacterial member to the growing family of plant cell wall degrading FAEs that at present is largely of fungal origin, and it benchmarks the bioproduction of FA from triticale bran.Peer reviewed: YesNRC publication: Ye

Preparative Asymmetric Reduction of Ketones in a Biphasic Medium with an (S)-AIcohol Dehydrogenase under in situ-Cofactor-Recycling with a Formate Dehydrogenase

Gröger H, Hummel W, Rollmann C, et al. Preparative Asymmetric Reduction of Ketones in a Biphasic Medium with an (S)-AIcohol Dehydrogenase under in situ-Cofactor-Recycling with a Formate Dehydrogenase. Tetrahedron. 2004;60(3):633-640

Discovery of a thermostable Baeyer–Villiger monooxygenase by genome mining

Baeyer–Villiger monooxygenases represent useful biocatalytic tools, as they can catalyze reactions which are difficult to achieve using chemical means. However, only a limited number of these atypical monooxygenases are available in recombinant form. Using a recently described protein sequence motif, a putative Baeyer–Villiger monooxygenase (BVMO) was identified in the genome of the thermophilic actinomycete Thermobifida fusca. Heterologous expression of the respective protein in Escherichia coli and subsequent enzyme characterization showed that it indeed represents a BVMO. The NADPH-dependent and FAD-containing monooxygenase is active with a wide range of aromatic ketones, while aliphatic substrates are also converted. The best substrate discovered so far is phenylacetone (kcat = 1.9 s−1, KM = 59 μM). The enzyme exhibits moderate enantioselectivity with α-methylphenylacetone (enantiomeric ratio of 7). In addition to Baeyer–Villiger reactions, the enzyme is able to perform sulfur oxidations. Different from all known BVMOs, this newly identified biocatalyst is relatively thermostable, displaying an activity half-life of 1 day at 52°C. This study demonstrates that, using effective annotation tools, genomes can efficiently be exploited as a source of novel BVMOs.