Expanding the set of rhodococcal Baeyer–Villiger monooxygenases by high-throughput cloning, expression and substrate screening

To expand the available set of Baeyer–Villiger monooxygenases (BVMOs), we have created expression constructs for producing 22 Type I BVMOs that are present in the genome of Rhodococcus jostii RHA1. Each BVMO has been probed with a large panel of potential substrates. Except for testing their substrate acceptance, also the enantioselectivity of some selected BVMOs was studied. The results provide insight into the biocatalytic potential of this collection of BVMOs and expand the biocatalytic repertoire known for BVMOs. This study also sheds light on the catalytic capacity of this large set of BVMOs that is present in this specific actinomycete. Furthermore, a comparative sequence analysis revealed a new BVMO-typifying sequence motif. This motif represents a useful tool for effective future genome mining efforts.

Mesozoic-Cenozoic crustaceans preserved within echinoid tests and bivalve shells

Associations of crustaceans with echinoids (Echinodermata) and bivalves (Mollusca) are not uncommon in modern oceans. Here we record the occurrence of anomurans, brachyurans and isopods within echinoid tests and bivalve shells from the Middle Jurassic of France, the Upper Jurassic of the Czech Republic, the Eocene of Croatia and the Miocene of Austria. Additionally a new genus and species of fossil cirolanid isopod from the Middle Jurassic of France is described. The present examples are interpreted as crustacean sheltering, probably for safe and undisturbed moulting (ecdysis), within a vacant host test or shell. However, accidental association (washed in) or even food remains cannot be ruled out entirelyWeb of Science90361160

The possible role of matrix metalloproteinase (MMP)-2 and MMP-9 in cancer, e.g. acute leukemia

In the past decades, a lot of effort has been put in identifying the role of matrix metalloproteinases (MMPs) in cancer. The main role of MMPs in angiogenesis, tumor growth and metastasis is degradation of extracellular matrix (ECM) and release and/or activation of growth factors through their degradative activity. The degradative activity finally results in cancer progression. MMP-inhibitors (MMPIs) have already been designed and tested, based on the degradative role of MMPs in cancer progression. First clinical trials with MMPIs have been performed with disappointing results, showing that in order to use MMP-inhibition the mechanisms underlying MMP-expression in cancer have to be further elucidated. This paper reviews the mechanisms of MMPs on molecular and cellular level and discusses the role for MMPs and MMP-inhibition in cancer with special focus on acute leukemia

Introducing an Artificial Deazaflavin Cofactor in Escherichia coli and Saccharomyces cerevisiae

[Image: see text] Deazaflavin-dependent whole-cell conversions in well-studied and industrially relevant microorganisms such as Escherichia coli and Saccharomyces cerevisiae have high potential for the biocatalytic production of valuable compounds. The artificial deazaflavin FOP (FO-5′-phosphate) can functionally substitute the natural deazaflavin F(420) and can be synthesized in fewer steps, offering a solution to the limited availability of the latter due to its complex (bio)synthesis. Herein we set out to produce FOP in vivo as a scalable FOP production method and as a means for FOP-mediated whole-cell conversions. Heterologous expression of the riboflavin kinase from Schizosaccharomyces pombe enabled in vivo phosphorylation of FO, which was supplied by either organic synthesis ex vivo, or by a coexpressed FO synthase in vivo, producing FOP in E. coli as well as in S. cerevisiae. Through combined approaches of enzyme engineering as well as optimization of expression systems and growth media, we further improved the in vivo FOP production in both organisms. The improved FOP production yield in E. coli is comparable to the F(420) yield of native F(420)-producing organisms such as Mycobacterium smegmatis, but the former can be achieved in a significantly shorter time frame. Our E. coli expression system has an estimated production rate of 0.078 μmol L(–1) h(–1) and results in an intracellular FOP concentration of about 40 μM, which is high enough to support catalysis. In fact, we demonstrate the successful FOP-mediated whole-cell conversion of ketoisophorone using E. coli cells. In S. cerevisiae, in vivo FOP production by SpRFK using supplied FO was improved through media optimization and enzyme engineering. Through structure-guided enzyme engineering, a SpRFK variant with 7-fold increased catalytic efficiency compared to the wild type was discovered. By using this variant in optimized media conditions, FOP production yield in S. cerevisiae was 20-fold increased compared to the very low initial yield of 0.24 ± 0.04 nmol per g dry biomass. The results show that bacterial and eukaryotic hosts can be engineered to produce the functional deazaflavin cofactor mimic FOP

Evolutionary bi-stability in pathogen transmission mode

Many pathogens transmit to new hosts by both infection (horizontal transmission) and transfer to the infected host's offspring (vertical transmission). These two transmission modes require speci®c adap- tations of the pathogen that can be mutually exclusive, resulting in a trade-off between horizontal and vertical transmission. We show that in mathematical models such trade-offs can lead to the simultaneous existence of two evolutionary stable states (evolutionary bi-stability) of allocation of resources to the two modes of transmission. We also show that jumping between evolutionary stable states can be induced by gradual environmental changes. Using quantitative PCR-based estimates of abundance in seed and vege- tative parts, we show that the pathogen of wheat, Phaeosphaeria nodorum, has jumped between two distinct states of transmission mode twice in the past 160 years, which, based on published evidence, we interpret as adaptation to environmental change. The ®nding of evolutionary bi-stability has impli- cations for human, animal and other plant diseases. An ill-judged change in a disease control programme could cause the pathogen to evolve a new, and possibly more damaging, combination of transmission modes. Similarly, environmental changes can shift the balance between transmission modes, with adverse effects on human, animal and plant health

Corrigendum to:Genome Mining of Oxidation Modules in trans -Acyltransferase Polyketide Synthases Reveals a Culturable Source for Lobatamides (Angewandte Chemie International Edition, (2020), 59, 20, (7761-7765), 10.1002/anie.201916005)

In the abstract and on page 7763 of this Communication, the authors erroneously describe Gynuella sunshinyii as the first culturable source for lobatamides. However, Suzumura et al. reported in 1997 the isolation of the compound YM-75518, which has the same structure as lobatamide A, from Pseudomonas sp. Q38009. This error does not affect the results of any of the experiments and data in this Communication. The authors sincerely apologize for this error

Discovery and characterization of a putrescine oxidase from Rhodococcus erythropolis NCIMB 11540

A gene encoding a putrescine oxidase (PuO(Rh), EC 1.4.3.10) was identified from the genome of Rhodococcus erythropolis NCIMB 11540. The gene was cloned in the pBAD vector and overexpressed at high levels in Escherichia coli. The purified enzyme was shown to be a soluble dimeric flavoprotein consisting of subunits of 50 kDa and contains non-covalently bound flavin adenine dinucleotide as a cofactor. From all substrates, the highest catalytic efficiency was found with putrescine (K(M) = 8.2 μM, k(cat) = 26 s(−1)). PuO(Rh) accepts longer polyamines, while short diamines and monoamines strongly inhibit activity. PuO(Rh) is a reasonably thermostable enzyme with t(1/2) at 50°C of 2 h. Based on the crystal structure of human monoamine oxidase B, we constructed a model structure of PuO(Rh), which hinted to a crucial role of Glu324 for substrate binding. Mutation of this residue resulted in a drastic drop (five orders of magnitude) in catalytic efficiency. Interestingly, the mutant enzyme showed activity with monoamines, which are not accepted by wt-PuO(Rh). ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s00253-007-1310-4) contains supplementary material, which is available to authorized users

Facile Stereoselective Reduction of Prochiral Ketones by using an F ₄₂₀-dependent alcohol dehydrogenase

Effective procedures for the synthesis of optically pure alcohols are highly valuable. A commonly employed method involves the biocatalytic reduction of prochiral ketones. This is typically achieved by using nicotinamide cofactor-dependent reductases. In this work, we demonstrate that a rather unexplored class of enzymes can also be used for this. We used an F420-dependent alcohol dehydrogenase (ADF) from Methanoculleus thermophilicus that was found to reduce various ketones to enantiopure alcohols. The respective (S) alcohols were obtained in excellent enantiopurity (>99 % ee). Furthermore, we discovered that the deazaflavoenzyme can be used as a self-sufficient system by merely using a sacrificial cosubstrate (isopropanol) and a catalytic amount of cofactor F420 or the unnatural cofactor FOP to achieve full conversion. This study reveals that deazaflavoenzymes complement the biocatalytic toolbox for enantioselective ketone reductions

Origin, early evolution and palaeoecology of Gymnopleura (Crustacea, Decapoda): Basal palaeocorystoid crabs from the Upper Jurassic-Lower Cretaceous of central Europe

Recent fieldwork has yielded new decapod crustacean material from Upper Jurassic-lowest Cretaceous bioclastic limestones at Kotou.c quarry near.Stramberk (Moravia, northeastern Czech Republic). Two specimens in this lot can be ascribed to the superfamily Palaeocorystoidea and represent the oldest gymnopleuran crabs known to date. A new genus and species, Moravacarcinus stramberkensis, are here erected and assigned to a new subfamily, Moravacarcininae, to accommodate this basal necrocarcinid. Our re-examination of Late Jurassic primitive crabs from southern Germany has resulted in the discovery of another early member of this group, here referred to a new genus, Juranecrocarcinus, as J. angulosum (Wehner, 1988). These new finds demonstrate that palaeocorystoids originated within shallow-water, reefal settings in the Upper Jurassic reef belt across central and southern Europe. We hypothesise that members of basal necrocarcinid subfamilies (Paranecrocarcininae and Moravacarcininae subfam. nov.), and thus the Gymnopleura, were derived from a dynomeniform ancestor which adapted to and became modified for a burying mode of life. Possible candidates are, for instance, the goniodromitid genus Cycloprosopon L.orenthey, in L.orenthey and Beurlen, 1929 and the longodromitid genera Longodromites Patrulius, 1959 and Planoprosopon Schweitzer, Feldmann and Laz.ar, 2007. The evolutionary pathways and palaeogeographical history of Mesozoic gymnopleurans were markedly influenced by the planktonic revolution which considerably enriched deeper-marine clastic sediments with nutrients from the Late Jurassic onwards.Web of Science564art. no. 11017