Title in bold letters

Title in bold letters

Enzymes and biotechnology: could we overcome modern challenges? 2nd International Conference on Genomics & Pharmacogenomics Dr. Junio Cota VTT Brasil LTDA 01/31/2020 SUMMARY What is VTT? The OMICS Era New Enzyme Discovery Protein Engineering

2 01/31/2020 3 VTT Group in brief Turnover 292 M (2010) Personnel 3,167 (1.1.2011) Customer sectors Focus areas of research Biotechnology, pharmaceutical Applied materials Bio- and chemical processes

Energy Information and communication and food industries Electronics Energy ICT Real estate and construction Machines and vehicles Services and logistics Forest industry Process industry and environment technologies Industrial systems management

Microtechnologies and electronics Services and the built environment Business research VTTs operations Research and Development Strategic Research Business Solutions IP Business Group Services VTTs companies VTT Expert Services Ltd (incl. Labtium Ltd, Enas Ltd) VTT Ventures Ltd VTT International Ltd VTT Memsfab Ltd 01/31/2020 4 01/31/2020

5 BIOREFINERY Brazilian Brazilianbiomass biomassraw rawmaterial material Process Sector Key service & offering Key

technologies End products Wood and fibre processing Biotechnical processing Energy production Pulp, Pulp,paper paper&&Packaging Packaging &&Solid wood

Solid wood Bio Bio&&Chemistry Chemistry Energy Energy Multi-technology Multi-technology solutions solutions&& pilot pilotscale scaleinfrastructure

infrastructure Biotechnical Biotechnical conversion conversion technologies technologies Thermal Thermalfuel fuel &&waste conversion waste conversion technologies

technologies Biomass Biomasshydrolysis, hydrolysis, Enzymes Enzymes&& Cell Cellfactories factories Gasification, Gasification,pyrolysis pyrolysis&& combustion combustiontechnologies

technologies Fibre Fibreprocessing, processing, Paper making, Paper making,Coating, Coating, Modelling & Simulation Modelling & Simulation Novel Novelfibre

fibrebased basedproducts products &&Advanced wood products Advanced wood products Chemicals, Chemicals, Bio Bioalcohols alcohols && Biomaterials Biomaterials

Biofuel, Biofuel,Electricity Electricity &&Heat Heat 01/31/2020 BRAZIL 6 01/31/2020 SUMMARY

What is VTT? The OMICS Era New Enzyme Discovery Protein Engineering 7 01/31/2020 8 Human Genome Project 1990 - 2003 http://web.ornl.gov/sci/techresources/Human_Genome/project/whydoe.shtml

01/31/2020 9 Evolution of Cost per Megabase http://evomics.org/2014/01/sequencing-technology-wheres-my-minion/ 01/31/2020 10 Evolution of Whole-Genome Sequencing Su, Andrew (2013): Cumulative sequenced genomes. figshare.

01/31/2020 11 Bioinformatics: a Modern Challenge Big Data, Genome Assembling, Gene Annotation, Computer Modeling 01/31/2020 SUMMARY What is VTT? The OMICS Era New Enzyme Discovery Protein Engineering

12 01/31/2020 13 Enzyme discovery in the OMICS Era Liebl, Wolfgang (2011): Metagenomics 01/31/2020 New Enzymes for Biofuels: GH 10 Xylanase 14

01/31/2020 New Enzymes for Biofuels: GH 10 Xylanase 15 01/31/2020 16 Proteomics: Secretome of Penicillium equinulatum 01/31/2020 17

Proteomics: Secretome of Penicillium equinulatum SCB: Sugar Cane Bagasse HDT: Hydrothermal Treatment SET: Steam Explosion Treatment SAT: Sulfuric Acid Treatment MCL: Microcrystalline Cellulose 01/31/2020 18 Proteomics: Secretome of Trichoderma harzianum 01/31/2020

19 Proteomics: Secretome of Trichoderma harzianum 01/31/2020 SUMMARY What is VTT? The OMICS Era New Enzyme Discovery Protein Engineering 20 01/31/2020

21 Protein Engineering is a tailor-made process Whats on your mind? Whats your need? 01/31/2020 22 Protein Engineering: Rational or Non-rational Design? Current Paradigms

Mechanism-based (Rational) Detailed structural analysis X Empiricism-based (Non-rational) Libraries based 01/31/2020 Building a Xylanase Lichenase Chimera 23

01/31/2020 24 Chimeras: Multidomain Proteins Multidomain/multifunctional proteins can reduce costs with enzyme load; End-to-end fusion between the N and C termini of the parental enzymes can result in nonfunctional chimeras. S.Y. Hong et al., Biotechnology Letters, 29, 931-936 (2007)

01/31/2020 25 Chimeras: Multidomain Proteins The selection of the linker sequence is particularly important for the construction of functional fusion proteins ? 01/31/2020

26 Building Chimeras: Molecular Dinamics Energy Landscape Theory Structure Based Models The topology could drives the protein folding

Save computational time X. L. Ji, and S. Q. Liu. J. Biomol Struct Dyn 28, 621-623 (2011) 01/31/2020 27 Structure Based Models (SB) The unique Free Energy basin suggests a group of structures candidates: simulations are mainly driven by the entropy of the system

01/31/2020 Building Chimera Overlap PCR SDS-PAGE 28 01/31/2020 29 SAXS experimental and theoretical curves Representative structure taken from

the Free Energy basin with 2=2.80, Rg=26.0 and CM distance=40.7 inset Theoretical scattering curve was generated in CRYSOL and the representation in VMD 01/31/2020 Optimal pH Xylan Parental enzyme Chimera

Lichenan 30 01/31/2020 Optimal Temperature Xylan Parental enzyme Chimera Lichenan

31 01/31/2020 Substrate Specificity Specific activity (U/nmol) Substrate Xylanase Lichenase XylLich Birchwood Xylan Beechwood Xylan

Rye Arabinoxylan 3.73 0.29 3.17 0.07 3.73 0.14 ND ND ND 2.71 0.13 2.87 0.08 3.03 0.15 Wheat Arabinoxylan Oat Spelt Xylan

1.36 0.12 3.28 0.27 ND ND 0.88 0.07 2.15 0.06 Lichenan -GlucanGlucan Laminarin ND ND

ND 3.65 0.29 5.03 0.20 ND 3.85 0.16 5.11 0.07 ND Xyloglucan Glucomannan (Konjac) ND ND

ND ND ND ND 32 01/31/2020 Capillary Electrophoresis Xylohexaose Lichenan

33 01/31/2020 Capillary Electrophoresis Xylohexaose + Lichenan 34 01/31/2020 Enzyme6 Yield times greater 20% greater

35 01/31/2020 36 Conclusions This work presented a novelty way to predict the disposal of chimera domains in solution before experimental assays; A potential tool for screening and development of enzyme

cocktails for second generation biofuels; The expansion of hydrolase activities in an unique protein could be a route for increase cost-effective of biomass saccharification; Enzyme production data suggests an advantage on producing the fused protein instead the wild type ones separated. 01/31/2020 37

Protein Engineering: Typical Challenges Design proteins with certain function; Design proteins which bind novel ligands; Alter binding affinity and specificity of proteins; Increase activity of enzymes; Change thermal tolerance, pH stability;

Decrease inhibition of enzymes; Alter allosteric regulation; Increase protease resistance; Reactivity in nonaqueous solvents; Eliminate cofactor requirement.

01/31/2020 38 Acknowledgements 01/31/2020 39 [email protected] [email protected]

Recently Viewed Presentations

  • La Rentrée des classes - Lakeridge Junior High School

    La Rentrée des classes - Lakeridge Junior High School

    Back to school in France ... = exams = tests/assessments = detentions = school reports = repeating a school year Le Cartable French children take a school bag with them, called a cartable Dans la trousse Things you might need...
  • Which Poem? Why? Ozymandias London The Prelude My

    Which Poem? Why? Ozymandias London The Prelude My

    The Emigrée (1993) . Carol . Rumens. There once was a country… I left it as a childbut my memory of it is sunlight-clearfor it seems I never saw it in that Novemberwhich, I am told, comes to the mildest...
  • Computer Architecture and Engineering Lecture 6: The Design ...

    Computer Architecture and Engineering Lecture 6: The Design ...

    Times New Roman Arial Symbol Courier New Monotype Corsiva Wingdings delay Autosketch Drawing Microsoft Visio Drawing CS152 Computer Architecture and Engineering Lecture 10 High-Level Design/ Microcode programming Recap: What's wrong with our CPI=1 processor?
  • UMBILICAL CORD STEM CELLS - University of Pittsburgh

    UMBILICAL CORD STEM CELLS - University of Pittsburgh

    Cellular vehicles for gene therapy Umbilical cord stem cells can be stably transduced using Retroviral vectors for necessary genetic correction and can be used for transplantation Future applications In autoimmune diseases, where there has been some success with autologous transplantation...
  • Shakespeare Sonnet 130 Analysis

    Shakespeare Sonnet 130 Analysis

    For your paraphrase, read the modern translation and then write a paragraph about telling what this sonnet is about. The speaker of the sonnet describes the woman he loves. He makes multiple comparisons of her to things like the sun,...
  • Thomas Paine's Common Sense - Mr. Cummings

    Thomas Paine's Common Sense - Mr. Cummings

    Thomas Paine's Common Sense Mr. Cummings US History ... Wrote another pamphlet against British monarchies - charged with seditious libel in Great Britain in 1792 - arrested on another charge in 1793 in France - after his release, fled to...
  • Safeguarding Forums for Schools Summer 2017

    Safeguarding Forums for Schools Summer 2017

    Working with Resistant Families and Disguised Compliance. Early Help. ESCB update. Southend, Essex and Thurrock (SET) agreed common approach to . CSE Champion role - following standards must be achieved:
  • Huzuraba's letter on 7th Jan, 2008

    Huzuraba's letter on 7th Jan, 2008

    Huzuraba's letter on finding faults with Jamaat 01-03-08 On the subject of errors, it has been brought to my attention that certain ladies are occupied with objectionable mannerism which is contrary to both the Jamaati and Islamic teachings. ... Sermon:...