Characterization of lignin-modifying enzymes of the selective white-rot fungus Physisporinus rivulosus

White-rot fungi are wood degrading organisms that are able to decompose all wood polymers; lignin, cellulose and hemicellulose. Especially the selective white-rot fungi that decompose preferentially wood lignin are promising for biopulping applications. In biopulping the pretreatment of wood chips with white-rot fungi enhances the subsequent pulping step and substantially reduces the refining energy consumption in mechanical pulping. Because it is not possible to carry out biopulping in industrial scale as a closed process it has been necessary to search for new selective strains of white-rot fungi which naturally occur in Finland and cause selective white-rot of Finnish wood raw-material. In a screening of 300 fungal strains a rare polypore, Physisporinus rivulosus strain T241i isolated from a forest burn research site, was found to be a selective lignin degrader and promising for the use in biopulping. Since selective lignin degradation is apparently essential for biopulping, knowledge on lignin-modifying enzymes and the regulation of their production by a biopulping fungus is needed. White-rot fungal enzymes that participate in lignin degradation are laccase, lignin peroxidase (LiP), manganese peroxidase (MnP), versatile peroxidase (VP) and hydrogen peroxide forming enzymes. In this study, P. rivulosus was observed to produce MnP, laccase and oxalic acid during growth on wood chips. In liquid cultures manganese and veratryl alcohol increased the production of acidic MnP isoforms detected also in wood chip cultures. Laccase production by P. rivulosus was low unless the cultures were supplemented with sawdust and charred wood, the components of natural growth environment of the fungus. In white-rot fungi the lignin-modifying enzymes are typically present as multiple isoforms. In this study, two MnP encoding genes, mnpA and mnpB, were cloned and characterized from P. rivulosus T241i. Analysis of the N-terminal amino acid sequences of two purified MnPs and putative amino acid sequence of the two cloned mnp genes suggested that P. rivulosus possesses at least four mnp genes. The genes mnpA and mnpB markedly differ from each other by the gene length, sequence and intron-exon structure. In addition, their expression is differentially affected by the addition of manganese and veratryl alcohol. P. rivulosus produced laccase as at least two isoforms. The results of this study revealed that the production of MnP and laccase was differentially regulated in P. rivulosus, which ensures the efficient lignin degradation under a variety of environmental conditions.Valkolahosienet ovat puuta lahottavia sieniä, jotka kykenevät hajottamaan kaikkia puun polymeerejä; ligniiniä, selluloosaa ja hemiselluloosaa. Valkolahoa aiheuttavien ligniiniä hajottavien sienten käyttöä puuhakkeiden käsittelyssä ennen massanvalmistusta nimitetään biopulppaukseksi. Erityisesti ns. selektiivisiä valkolahosieniä eli sieniä, jotka hajottavat puuaineesta suhteellisesti enemmän ligniiniä kuin muita komponentteja, on pidetty lupaavina biopulppaussieninä. Koska biopulppausta ei teollisessa mittakaavassa ole mahdollista toteuttaa suljettuna prosessina, on ollut tarpeellista etsiä ja tutkia tähän tarkoitukseen soveltuvia Suomessa luonnonvaraisena esiintyviä sienikantoja. Noin 300 sienen vertailututkimuksessa erittäin harvinainen metsäpaloalueilla esiintyvä talikääpä, Physisporinus rivulosus kanta T241i, osoittautui selektiiviseksi ligniininhajottajaksi ja biopulppausominaisuuksiltaan lupaavaksi. Ligniinin hajotukseen osallistuvia entsyymejä ovat lakkaasi, ligniiniperoksidaasi (LiP), mangaaniperoksidaasi (MnP), hybridityyppinen peroksidaasi (versatile peroxidase; VP) sekä vetyperoksidia tuottavat entsyymit. Tässä työssä P. rivulosus T241i:n havaittiin erittävän puuhakkeella kasvaessaan oksaalihappoa sekä MnP:a ja lakkaasia. Mangaani ja veratryylialkoholi edistivät liuosviljelmässä MnP:n tuottoa. Lakkaasia P. rivulosus tuotti liuosviljelmässä melko vähän, jos alustassa ei ollut sen luontaiseen kasvuympäristöön kuuluvia kiinteitä substraatteja. Valkolahosienille on tyypillistä, että niiden ligniiniin vaikuttavat entsyymit esiintyvät lukuisina isomuotoina. P. rivulosus T241i:lta karakterisoitiin kaksi eri MnP:a koodaavaa geeniä, mnpA ja mnpB, mutta MnP:n N-terminaalisten aminohapposekvenssien analyysin perusteella on todennäköistä, että sienellä on mnpA:n ja mnpB:n lisäksi ainakin kaksi muuta mnp-geeniä. Geenit mnpA ja mnpB poikkesivat merkittävästi toisistaan sekä geenin pituuden että introni-eksoni -rakenteen suhteen. Lisäksi veratryylialkoholi ja mangaani vaikuttivat P. rivulosusin mnpA ja mnpB geenien ilmentymiseen eri tavalla. P. rivulosusista löydettiin kaksi lakkaasi-isomuotoa. Tämän tutkimuksen tulosten mukaan P. rivulosus:lla erot kasvuolosuhteissa vaikuttivat eri entsyymien eritykseen eri tavalla, mikä takaa sienen tehokkaan ligniinin hajotuksen erilaisissa kasvuolosuhteissa

Biotechnology as a competitive edge for the Finnish forest cluster

In this study we have collected information by interviewing all identified parties within the Finnish forest sector who might have a potential biotechnology connection : university research groups, research institutions, small and medium-sized biotechnology-companies and up to the largest forest companies. The ultimate goal was to assess how resources have been allocated and biotechnologies utilized within the value chain of the entire forest sector. This study aimed at providing answers to the following questions : • What are the current Finnish academic resources and projects related to forest industry biotechnology? • How much does the Finnish forest cluster invest in biotechnology R&D activity, and what are the key application areas in the value chain? • How well do the academic resources, company R&D investments and research needs converge to help secure the future competitiveness of the Finnish forest industries? In order to answer the questions above, the study approached the matter in consecutive steps. First, the existing forest industry related biotechnological knowledge base within the academia on one hand, and the resource base among firms on the other hand, were mapped. Following up on that, we evaluated the sales expectations of forestry related biotechnological applications within the domestic forestry cluster itself, other potential domestic industries and global export markets. The third step assessed whether the development of forestry related biotechnological applications is justifiable in the framework of comparative advantage. This was accomplished by comparing the relevant existing knowledge and other resource bases to their sales expectations. In order to evaluate the potential of biotechnology in the entire forest industry value chain, the study assessed four value chain modules. Module 1 represents the beginning of the value chain : forestry applications. Module 2 consists of the development of wood products, module 3 is related to the pulp and paper industry, and module 4 to utilization side streams for bioenergy, biochemicals and other food or pharmaceutical applications. The assessment of module 1 implies that there is a constant lack of resources. Basic research conducts some relatively long projects, which often seem too time-consuming in applied research and corporate R&D. There seems to be only few active links between the academic research projects and companies. Many new technologies already exist but since the individual forest owners hardly have incentives to invest in R&D due to e.g. the long breeding cycle, collaboration with companies seems as the only potential pathway to commercialization of forestry related biotechnologies. There were few biotechnology-based projects within the module 2. The research and product development seems to focus on physical modifications, and composite research is based on chemistry. Module 3, paper, pulp and board industry, seems to be the most active in research and product development activity. Their products generate positive cash flows, and research projects are abundantly funded. The companies are closely involved in the research projects as financiers and collaborators. This involvement impacts on the nature of the research, which seems highly applicable and linked closely to industrial applications. Consequently, biotechnology applications are already used in the pulp and paper industry. Some biotechnology applications are adopted rapidly. They, such as enzymes in reducing paper machine runnability problems, do not affect the quality of the fibers, intermediate or end products and are thus easier to take into use in production scale. We observed the research and product development within module 4 as a high priority for both the academia and industry. The research is anticipated to grow strongly and even more than in other modules. Biotechnologies are applied as substitutes to chemical and thermal technologies. However, all of these fields of technology are developed and applied by the industry. This provides some important implications for technology development and innovation policy. Due to the fuzziness between technology border-lines, it seems misleading to prioritize biotechnologies over some other technology; in contrast, the most efficient technology should be preferred. Accordingly, technology subsidies might be most efficient if the public technology programmes would be based on application segments instead of a specific technology. Our assessment of international patenting activity raised some interesting notions. Finland seemed to be comparatively most specialized in plant genetic engineering, food and food additive, and waste disposal and the environment applications. However, biotechnology based biofuels are not included as a source of comparative advantage, which also stresses the importance of parallel development of biotechnologies and other technology fields. A potential source of value creation could be the utilization of process side streams more efficiently, including refinement of by-products such as tall oil, to products with higher value added in other application areas.The paper and board making might also be strongly influenced by new packaging solutions, materials and methods; these utilize, however, only rarely or never biotechnologies as such. Finland has a good overall and mainly publicly maintained infrastructure. If the raw material’s high quality and some special features can compensate the relatively low growth rates, Finland should be able to attract the multinational pulp and paper industry also in the long term. We conclude that the development of biotechnologies should not contain any intrinsic value per se. The commercial value of the biotechnology could be benchmarked with the value of alternative technologies; and consequently, biotechnology could become part of the technology options for companies active in established and conventional industries. The Finnish forest cluster has financial resources to commercialize any new technology that can increase the process efficiency or provide other economic benefits in new application areas. This is a reason why we see this area exceptionally promising compared to any other high technology field without such a financial backbone

Kommunens namn i kommunsloganer : perspektiv från lingvistik och marknadsföring

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Visual performance with small concave and convex displays

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A frameshift mutation in ARMC3 is associated with a tail stump sperm defect in Swedish Red (Bos taurus) cattle

Background: Artificial insemination is widely used in many cattle breeding programs. Semen samples of breeding bulls are collected and closely examined immediately after collection at artificial insemination centers. Only ejaculates without anomalous findings are retained for artificial insemination. Although morphological aberrations of the spermatozoa are a frequent reason for discarding ejaculates, the genetic determinants underlying poor semen quality are scarcely understood. Results: A tail stump sperm defect was observed in three bulls of the Swedish Red cattle breed. The spermatozoa of affected bulls were immotile because of severely disorganized tails indicating disturbed spermatogenesis. We genotyped three affected bulls and 18 unaffected male half-sibs at 46,035 SNPs and performed homozygosity mapping to map the fertility disorder to an 8.42 Mb interval on bovine chromosome 13. The analysis of whole-genome re-sequencing data of an affected bull and 300 unaffected animals from eleven cattle breeds other than Swedish Red revealed a 1 bp deletion (Chr13: 24,301,425 bp, ss1815612719) in the eleventh exon of the armadillo repeat containing 3-encoding gene (ARMC3) that was compatible with the supposed recessive mode of inheritance. The deletion is expected to alter the reading frame and to induce premature translation termination (p.A451fs26). The mutated protein is shortened by 401 amino acids (46 %) and lacks domains that are likely essential for normal protein function. Conclusions: We report the phenotypic and genetic characterization of a sterilizing tail stump sperm defect in the Swedish Red cattle breed. Exploiting high-density genotypes and massive re-sequencing data enabled us to identify the most likely causal mutation for the fertility disorder in bovine ARMC3. Our results provide the basis for monitoring the mutated variant in the Swedish Red cattle population and for the early identification of infertile animals.Peer reviewe

Measurement of t(t)over-bar normalised multi-differential cross sections in pp collisions at root s=13 TeV, and simultaneous determination of the strong coupling strength, top quark pole mass, and parton distribution functions

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An embedding technique to determine ττ backgrounds in proton-proton collision data

An embedding technique is presented to estimate standard model tau tau backgrounds from data with minimal simulation input. In the data, the muons are removed from reconstructed mu mu events and replaced with simulated tau leptons with the same kinematic properties. In this way, a set of hybrid events is obtained that does not rely on simulation except for the decay of the tau leptons. The challenges in describing the underlying event or the production of associated jets in the simulation are avoided. The technique described in this paper was developed for CMS. Its validation and the inherent uncertainties are also discussed. The demonstration of the performance of the technique is based on a sample of proton-proton collisions collected by CMS in 2017 at root s = 13 TeV corresponding to an integrated luminosity of 41.5 fb(-1).Peer reviewe

Measurement of the Splitting Function in &ITpp &ITand Pb-Pb Collisions at root&ITsNN&IT=5.02 TeV

Data from heavy ion collisions suggest that the evolution of a parton shower is modified by interactions with the color charges in the dense partonic medium created in these collisions, but it is not known where in the shower evolution the modifications occur. The momentum ratio of the two leading partons, resolved as subjets, provides information about the parton shower evolution. This substructure observable, known as the splitting function, reflects the process of a parton splitting into two other partons and has been measured for jets with transverse momentum between 140 and 500 GeV, in pp and PbPb collisions at a center-of-mass energy of 5.02 TeV per nucleon pair. In central PbPb collisions, the splitting function indicates a more unbalanced momentum ratio, compared to peripheral PbPb and pp collisions.. The measurements are compared to various predictions from event generators and analytical calculations.Peer reviewe

Search for Physics beyond the Standard Model in Events with Overlapping Photons and Jets

Results are reported from a search for new particles that decay into a photon and two gluons, in events with jets. Novel jet substructure techniques are developed that allow photons to be identified in an environment densely populated with hadrons. The analyzed proton-proton collision data were collected by the CMS experiment at the LHC, in 2016 at root s = 13 TeV, and correspond to an integrated luminosity of 35.9 fb(-1). The spectra of total transverse hadronic energy of candidate events are examined for deviations from the standard model predictions. No statistically significant excess is observed over the expected background. The first cross section limits on new physics processes resulting in such events are set. The results are interpreted as upper limits on the rate of gluino pair production, utilizing a simplified stealth supersymmetry model. The excluded gluino masses extend up to 1.7 TeV, for a neutralino mass of 200 GeV and exceed previous mass constraints set by analyses targeting events with isolated photons.Peer reviewe

Electroweak production of two jets in association with a Z boson in proton-proton collisions root s =13 TeV

A measurement of the electroweak (EW) production of two jets in association with a Z boson in proton-proton collisions at root s = 13 TeV is presented, based on data recorded in 2016 by the CMS experiment at the LHC corresponding to an integrated luminosity of 35.9 fb(-1). The measurement is performed in the lljj final state with l including electrons and muons, and the jets j corresponding to the quarks produced in the hard interaction. The measured cross section in a kinematic region defined by invariant masses m(ll) > 50 GeV, m(jj) > 120 GeV, and transverse momenta P-Tj > 25 GeV is sigma(EW) (lljj) = 534 +/- 20 (stat) fb (syst) fb, in agreement with leading-order standard model predictions. The final state is also used to perform a search for anomalous trilinear gauge couplings. No evidence is found and limits on anomalous trilinear gauge couplings associated with dimension-six operators are given in the framework of an effective field theory. The corresponding 95% confidence level intervals are -2.6 <cwww/Lambda(2) <2.6 TeV-2 and -8.4 <cw/Lambda(2) <10.1 TeV-2. The additional jet activity of events in a signal-enriched region is also studied, and the measurements are in agreement with predictions.Peer reviewe