DOCSLIB.ORG

(12) United States Patent (10) Patent No.: US 9,695.454 B2 Natunen Et Al

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
(12) United States Patent (10) Patent No.: US 9,695.454 B2 Natunen Et Al USOO9695454B2 (12) United States Patent (10) Patent No.: US 9,695.454 B2 Natunen et al. (45) Date of Patent: Jul. 4, 2017 (54) PRODUCTION OF FUCOSYLATED (56) References Cited GLYCOPROTEINS U.S. PATENT DOCUMENTS (71) Applicants: Glykos Finland OY, Helsinki (FI); Novartis AG, Basel (CH) 4,758,512 A 7/1988 Goldberg et al. 5,116,964 A 5/1992 Capon et al. (72) Inventors: Jari Natunen, Vantaa (FI); Ann 5,428, 130 A 6/1995 Capon et al. Westerholm-Parvinen, Kirkkonummi 5,455,165 A 10/1995 Capon et al. (FI); Hanna Salo, Helsinki (FI); Jussi 5,514,582 A 5/1996 Capon et al. 5,674,728 A 10, 1997 Buxton et al. Joensuu, Tervakoski (FI); Heli Viskari, 5,693,520 A 12/1997 Branner et al. Nummela (FI); Christopher 5,756.338 A 5/1998 Buxton et al. Landowski, Helsinki (FI); Anne 5,776,730 A 7, 1998 Stuart Huuskonen, Helsinki (FI); Anne 5,821, 104 A 10, 1998 Holm et al. 5,840,570 A 11/1998 Berka et al. Kanerva, Helsinki (FI); Anna-Liisa 5,846,802 A 12/1998 Buxton et al. Hanninen, Helsinki (FI); Noora 5,968,774 A 10, 1999 Lehmbeck Salovuori, Helsinki (FI); Merja 5,989,889 A 11/1999 Rey et al. Penttila, Helsinki (FI); Juhani 6,013,452 A 1/2000 Christensen et al. Saarinen, Helsinki (FI); Markku 6,013,489 A 1/2000 Musters et al. 6,025, 185 A 2/2000 Christensen et al. Saloheimo, Helsinki (FI) 6,291,209 B1 9, 2001 Lehmbeck (73) Assignee: GLYKOS FINLAND OY, Helsinki 6,352,841 B1 3/2002 Lehmbeck (FI) 6,509,171 B1 1/2003 Berka et al. (*) Notice: Subject to any disclaimer, the term of this (Continued) patent is extended or adjusted under 35 U.S.C. 154(b) by 284 days. FOREIGN PATENT DOCUMENTS Appl. No.: EP 1266 011 B1 5, 2010 (21) 14/402,837 WO 96,36718 A1 11, 1996 (22) PCT Fed: May 23, 2013 WO 97.12045 A1 4f1997 WO 97,46689 A1 12/1997 (86) PCT No.: PCT/EP2013/060627 WO OOf 46375 A2 8, 2000 WO 2004/067709 A2 8, 2004 S 371 (c)(1), WO 2005/055944 A2 6, 2005 (2) Date: Nov. 21, 2014 WO 2005/087922 A1 9, 2005 PCT Pub. No.: WO2013/174927 WO 2008/07.4499 A1 6, 2008 (87) WO 2008, 112092 A2 9, 2008 PCT Pub. Date: Nov. 28, 2013 WO 2010/036898 A1 4/2010 WO 2011/07.5677 A2 6, 2011 (65) Prior Publication Data WO 2011/106389 A1 9, 2011 US 2015/O176044 A1 Jun. 25, 2015 OTHER PUBLICATIONS Related U.S. Application Data Ahamed, et al. Chymostatin can combine with pepstatin to eliminate Provisional application No. 61/650,910, filed on May extracellular protease activity in cultures of Aspergillus niger (60) NRRL-3. J. Ind. Microbiol. Biotechnol. 34: 165-169 (2007). 23, 2012. Baldwin, et al. Develop Systems for Manufacturing 100,000,000 (51) Int. C. Doses of an Emergency Pharmaceutical (e.g. Vaccine or Monoclo CI2R L/885 (2006.01) nal Antibody) Within 2 Months of Product Identification, Genencor CI2P 2L/00 (2006.01) International (Jun. 6, 2009). C07K I4/435 (2006.01) (Continued) CI2N 9/04 (2006.01) CI2N 9/10 (2006.01) CI2N 9/58 (2006.01) Primary Examiner — Karen Cochrane Carlson CI2N 9/88 (2006.01) (74) Attorney, Agent, or Firm — Patrick J. Halloran CI2N 15/52 (2006.01) C07K 6/00 (2006.01) CI2N 15/80 (2006.01) (57) ABSTRACT (52) U.S. C. Described herein are compositions including filamentous CPC ...... CI2P21/005 (2013.01); C07K 14/43545 fungal cells, such as Trichoderma fungal cells, having (2013.01); C07K 16/00 (2013.01): CI2N reduced protease activity and expressing fucosylation path 9/0006 (2013.01); C12N 9/1051 (2013.01); way. Further described herein are methods for producing a CI2N 9/58 (2013.01); CI2N 9/88 (2013.01); glycoprotein having fucosylated N-glycan, using genetically CI2N 15/52 (2013.01); C12N 15/80 modified filamentous fungal cells, for example, (2013.01); C07K 2319/01 (2013.01) Trichoderma fungal cells, as the expression system. (58) Field of Classification Search CPC ................................. C12R 1/885; C12P 21/15 See application file for complete search history. 17 Claims, 21 Drawing Sheets US 9,695.454 B2 Page 2 (56) References Cited 2012/0276075 A1 11, 2012 Monod et al. 2012/0288892 Al 11/2012 Maiyuran et al. U.S. PATENT DOCUMENTS 2012/0328626 A1 12/2012 Sethuraman et al. 2013 (OO 11875 A1 1/2013 Meehl et al. 6,602,684 B1 8, 2003 Umaha et al. 2014/0212977 A1 7/2014 Yaver et al. 6,806,062 B1 10, 2004 Hjort et al. 6,946,292 B2 9, 2005 Kanda et al. OTHER PUBLICATIONS 7,029,872 B2 4, 2006 Gerngross 7,094,530 B1 8, 2006 Sasaki et al. 7,122,330 B2 10, 2006 Emalfarb et al. Hintz et al., Improved gene expression in Aspergillus nidulans. Can. 7,163,804 B1 1/2007 Royer et al. Jo. Bot. 73 (Supp. 1): S876-S884 (1995). 7,198.938 B2 4, 2007 Shuster et al. Idiris, et al. Enhanced protein secretion from multiprotease-deficient 7,303,877 B2 12, 2007 Connelly et al. fission yeast by modification of its vacuolar protein sorting pathway. 7,323,327 B2 1, 2008 Edens et al. Appl. Microbiol. Biotechnol. 85: 667-677 (2010). 7,691,621 B2 4, 2010 Wang 7,771,971 B2 8, 2010 Connelly et al. Krysan, et al. Yapsins are a family of aspartyl proteases required for 7,794,974 B2 9, 2010 Pei et al. cell wall integrity in Saccharomyces cerevisiae. Eukaryotic Cell. 7,858,360 B2 12, 2010 Hansen et al. 4(8): 1364-1374 (2005). 7968,312 B2 6, 2011 Sagt et al. Kuroda, et al. Antibody expression in protease-deficient strains of 7,977,067 B2 T 2011 Power et al. the methlotrophic yeast ogataea minuta. FEMS Yeast Res. 7: 8,017,341 B2 9, 2011 Nikolaev et al. 1307-1316 (2007). 8,119,171 B2 2, 2012 Lopez et al. Martinez, et al. Genome sequencing and analysis of the biomass 8,288,517 B2 10, 2012 Clarkson et al. 8,389,269 B2 3/2013 Sagt et al. degrading fungus Trichoderma reesei (syn. Hypocrea jecorina). 8,426,164 B2 4, 2013 Hjort et al. Nature Biotech. 26(5): 553-560 (2008). 8.450,098 B2 5, 2013 Kim et al. Martinez, et al. Uniprot GORG34, XP002712642 (Oct. 19, 2011). 8,633,010 B2 1, 2014 Lehmbeck et al. Martinez, et al. Uniprot GORIW3, XP002712643 (Oct. 19, 2011). 8,647,856 B2 2, 2014 Shasky et al. Martinez, et al. Uniprot GORHO5, XP002712644 (Oct. 19, 2011). 8,680,252 B2 3, 2014 Emalfarb et al. Martinez, et al. Uniprot GORSP8, XP002712645 (Oct. 19, 2011). 8,716,004 B2 5, 2014 Wang 8,741,654 B2 6, 2014 Bodie et al. Martinez, et al. Uniprot GORVKO, XP002712646 (Oct. 19, 2011). 8,812.247 B2 8, 2014 Roubos et al. Martinez, et al. Uniprot GOR8TO, XP002712647 (Oct. 19, 2011). 8,916,363 B2 12, 2014 Gusakov et al. Van Den Hobergh, et al. Disruption of three acid proteases in 8,986,974 B2 3/2015 Maiyuran ................ C12N 9,58 Aspergillus niger. Eur, J. Biochem. 247: 605-613 (1997). 435/254.1 Yoon, et al. Construction of quintuple protease gene disruptant for 2002fOO58313 A1 5, 2002 Renkonen et al. heterologous protein production in aspergillus oryzae Appl. 2002O132320 A1 9, 2002 Wang et al. 2003. O157108 A1 8, 2003 Presta Microbiol. Biotechnol. 82: 691-701 (2009). 2003/0175884 A1 9, 2003 Umana et al. Yoon, et al. Disruption of ten protease genes in the filamentous 2004/0014170 A1 1, 2004 Miura et al. fungus aspergillus oryzae highly improves production of 2004/OO18573 A1 1, 2004 Power et al. heterologous proteins. Appl. Microbiol. Biotechnol. (Oct. 19, 2010). 2004/OO18590 A1 1, 2004 Gerngross et al. Adav et al., “Proteomic Analysis of pH and Strains Dependent 2004/OO72290 A1 4, 2004 Umana et al. Protein Secretion of Trichoderma Reesei’, J Proteome Res., (10) 10, 2004/00936.21 A1 5, 2004 Shitara et al. Oct. 7, 2011, pp. 4579-4596. 2004/O136986 A1 T/2004 Raju Archer et al., “Proteolytic Degradation of Heterologous Proteins 2004/0171826 A1 9, 2004 Hamilton 2004/019 1256 A1 9, 2004 Raju Expressed in Aspergillus Niger. Biotechnology Letters, vol. 14. 2004/0230.042 A1 11, 2004 Hamilton Issue 5, May 5, 1992, pp. 357-362. 2004/0241817 A1 12, 2004 Umana et al. Behnsen et al., “Secreted Aspergillus Fumigatus Protease Alp1 2004/O259150 A1 12, 2004 Nakamura et al. Degrades Human Complement Proteins C3, C4, and C5'. Infect 2005/0170452 A1 8, 2005 Wildt et al. Immun., 78(8), Aug. 2010, pp. 3585-3594. 2005/02O8617 A1 9, 2005 Bobrowicz et al. Berka et al., “Molecular Cloning and Deletion of the Gene Encoding 2005/0276805 A1 12, 2005 Hanai et al. Aspergillopepsin a from Aspergillus Awamori'. Gene..., 86(2), Feb. 2005/02771.54 A1 12, 2005 Dukler et al. 14, 1990, pp. 153-162. 2006, OOOO996 A1 1, 2006 Soldo Broekhuijsen et al., “Secretion of Heterologous Proteins by 2006/0040353 A1 2, 2006 Davidson et al. 2006/016O179 A1 T/2006 Bobrowicz et al. Aspergillus Niger: Production of Active Human Interleukin-6 in a 2006/0211085 A1 9, 2006 Bobrowicz Protease-Deficient Mutant by KEX2-like Processing of a 2006/0234345 A1 10, 2006 Schwartz et al. Glucoamylase-hIL6 Fusion Protein'. Journal of Biotechnology, 2006/0286637 A1 12, 2006 Hamilton 31(2), Nov.
Load more

Recommended publications
  • WO 2013/102674 A2 11 July 2013 (11.07.2013) W P O P C T
    (12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization International Bureau (10) International Publication Number (43) International Publication Date WO 2013/102674 A2 11 July 2013 (11.07.2013) W P O P C T (51) International Patent Classification: (74) Agent: CABINET PLASSERAUD; 235 cours Lafayette, C12N 1/15 (2006.01) F-69006 Lyon (FR). (21) International Application Number: (81) Designated States (unless otherwise indicated, for every PCT/EP20 13/050 126 kind of national protection available): AE, AG, AL, AM, AO, AT, AU, AZ, BA, BB, BG, BH, BN, BR, BW, BY, (22) Date: International Filing BZ, CA, CH, CL, CN, CO, CR, CU, CZ, DE, DK, DM, 4 January 2013 (04.01 .2013) DO, DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, (25) Filing Language: English HN, HR, HU, ID, IL, IN, IS, JP, KE, KG, KM, KN, KP, KR, KZ, LA, LC, LK, LR, LS, LT, LU, LY, MA, MD, (26) Publication Language: English ME, MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, (30) Priority Data: NO, NZ, OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, 61/583,559 5 January 2012 (05.01 .2012) US RW, SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, (71) Applicants: NOVARTIS INTERNATIONAL PHAR¬ ZM, ZW. MACEUTICAL LTD.; 13 1 Front Street, Hamilton (BM). GLYKOS FINLAND OY [FI/FI]; Viikinkaari 6, FI- (84) Designated States (unless otherwise indicated, for every 00790 Helsinki (FI).
    [Show full text]
  • (12) Patent Application Publication (10) Pub. No.: US 2016/0346364 A1 BRUNS Et Al
    US 2016.0346364A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2016/0346364 A1 BRUNS et al. (43) Pub. Date: Dec. 1, 2016 (54) MEDICAMENT AND METHOD FOR (52) U.S. Cl. TREATING INNATE IMMUNE RESPONSE CPC ........... A61K 38/488 (2013.01); A61K 38/482 DISEASES (2013.01); C12Y 304/23019 (2013.01); C12Y 304/21026 (2013.01); C12Y 304/23018 (71) Applicant: DSM IPASSETS B.V., Heerlen (NL) (2013.01); A61K 9/0053 (2013.01); C12N 9/62 (2013.01); A23L 29/06 (2016.08); A2ID 8/042 (72) Inventors: Maaike Johanna BRUINS, Kaiseraugst (2013.01); A23L 5/25 (2016.08); A23V (CH); Luppo EDENS, Kaiseraugst 2002/00 (2013.01) (CH); Lenneke NAN, Kaiseraugst (CH) (57) ABSTRACT (21) Appl. No.: 15/101,630 This invention relates to a medicament or a dietary Supple (22) PCT Filed: Dec. 11, 2014 ment comprising the Aspergillus niger aspergilloglutamic peptidase that is capable of hydrolyzing plant food allergens, (86). PCT No.: PCT/EP2014/077355 and more particularly, alpha-amylase/trypsin inhibitors, thereby treating diseases due to an innate immune response S 371 (c)(1), in humans, and/or allowing to delay the onset of said (2) Date: Jun. 3, 2016 diseases. The present invention relates to the discovery that (30) Foreign Application Priority Data the Aspergillus niger aspergilloglutamic peptidase is capable of hydrolyzing alpha-amylase/trypsin inhibitors that are Dec. 11, 2013 (EP) .................................. 13196580.8 present in wheat and related cereals said inhibitors being strong inducers of innate immune response. Furthermore, Publication Classification the present invention relates to a method for hydrolyzing alpha-amylase/trypsin inhibitors comprising incubating a (51) Int.
    [Show full text]
  • Proteolytic Enzymes in Grass Pollen and Their Relationship to Allergenic Proteins
    Proteolytic Enzymes in Grass Pollen and their Relationship to Allergenic Proteins By Rohit G. Saldanha A thesis submitted in fulfilment of the requirements for the degree of Masters by Research Faculty of Medicine The University of New South Wales March 2005 TABLE OF CONTENTS TABLE OF CONTENTS 1 LIST OF FIGURES 6 LIST OF TABLES 8 LIST OF TABLES 8 ABBREVIATIONS 8 ACKNOWLEDGEMENTS 11 PUBLISHED WORK FROM THIS THESIS 12 ABSTRACT 13 1. ASTHMA AND SENSITISATION IN ALLERGIC DISEASES 14 1.1 Defining Asthma and its Clinical Presentation 14 1.2 Inflammatory Responses in Asthma 15 1.2.1 The Early Phase Response 15 1.2.2 The Late Phase Reaction 16 1.3 Effects of Airway Inflammation 16 1.3.1 Respiratory Epithelium 16 1.3.2 Airway Remodelling 17 1.4 Classification of Asthma 18 1.4.1 Extrinsic Asthma 19 1.4.2 Intrinsic Asthma 19 1.5 Prevalence of Asthma 20 1.6 Immunological Sensitisation 22 1.7 Antigen Presentation and development of T cell Responses. 22 1.8 Factors Influencing T cell Activation Responses 25 1.8.1 Co-Stimulatory Interactions 25 1.8.2 Cognate Cellular Interactions 26 1.8.3 Soluble Pro-inflammatory Factors 26 1.9 Intracellular Signalling Mechanisms Regulating T cell Differentiation 30 2 POLLEN ALLERGENS AND THEIR RELATIONSHIP TO PROTEOLYTIC ENZYMES 33 1 2.1 The Role of Pollen Allergens in Asthma 33 2.2 Environmental Factors influencing Pollen Exposure 33 2.3 Classification of Pollen Sources 35 2.3.1 Taxonomy of Pollen Sources 35 2.3.2 Cross-Reactivity between different Pollen Allergens 40 2.4 Classification of Pollen Allergens 41 2.4.1
    [Show full text]
  • Review Article the Role of Microbial Aspartic Protease Enzyme in Food and Beverage Industries
    Hindawi Journal of Food Quality Volume 2018, Article ID 7957269, 15 pages https://doi.org/10.1155/2018/7957269 Review Article The Role of Microbial Aspartic Protease Enzyme in Food and Beverage Industries Jermen Mamo and Fassil Assefa Microbial, Cellular and Molecular Biology Department, College of Natural Science, Addis Ababa University, P.O. Box 1176, Addis Ababa, Ethiopia Correspondence should be addressed to Jermen Mamo; [email protected] Received 3 April 2018; Revised 16 May 2018; Accepted 29 May 2018; Published 3 July 2018 Academic Editor: Antimo Di Maro Copyright © 2018 Jermen Mamo and Fassil Assefa. is is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Proteases represent one of the three largest groups of industrial enzymes and account for about 60% of the total global enzymes sale. According to the Nomenclature Committee of the International Union of Biochemistry and Molecular Biology, proteases are classied in enzymes of class 3, the hydrolases, and the subclass 3.4, the peptide hydrolases or peptidase. Proteases are generally grouped into two main classes based on their site of action, that is, exopeptidases and endopeptidases. Protease has also been grouped into four classes based on their catalytic action: aspartic, cysteine, metallo, and serine proteases. However, lately, three new systems have been dened: the threonine-based proteasome system, the glutamate-glutamine system of eqolisin, and the serine-glutamate-aspartate system of sedolisin. Aspartic proteases (EC 3.4.23) are peptidases that display various activities and specicities.
    [Show full text]
  • Molecular Modelling for Enzyme-Inhibition: a Search for a New Treatment for Cataract and New Antimicrobials and Herbicides
    Molecular Modelling for Enzyme-Inhibition: A Search for a New Treatment for Cataract and New Antimicrobials and Herbicides A thesis submitted in partial fulfilment of the requirements for the Degree of Doctor of Philosophy in Biochemistry at the University of Canterbury Christchurch New Zealand Blair Gibb Stuart March 2010 Contents CONTENTS ACKNOWLEDGEMENTS 1 ABSTRACT AND PUBLISHED WORK 2 1 INTRODUCTION 6 1.1 Calpain and the cataract hypothesis 6 1.2 Proteases 7 1.3 Calpains 10 1.4 Structure of the eye, cataract and the importance of an anti-cataract drug 14 1.5 The β-strand: important for protease recognition 15 1.6 Computer modelling programs 17 1.7 References 20 2 DEVELOPMENT OF A CALPAIN MODEL FOR DOCKING STUDIES 27 2.1 Introduction 27 2.1.1 Overview of calpain model development 27 2.2 Calpain X-ray crystal structures 28 2.2.1 The first published structures 28 2.2.2 Calpain constructs 1KXR and 1MDW 30 2.3 Exploring the calpain construct 1KXR to develop a viable model for Glide docking experiments 33 2.4 The InducedFit docking model 37 Contents 2.5 Conclusion 37 2.6 References 39 3 MOLECULAR MODELING OF ACYCLIC INHIBITORS 43 3.1 Introduction 43 3.1.1 Natural inhibitors 43 3.1.2 Modified natural inhibitors 45 3.1.3 Lead compound: SJA-6017 46 3.2 Docking studies of known inhibitors 46 3.2.1 Compounds of Inoue et al 46 3.2.2 Docking results for the Inoue et al compounds 50 3.3 Docking Studies of SJA-6017 analogues 58 3.3.1 N-Heterocyclic dipeptides 58 3.3.2 Docking results of N-heterocyclic dipeptides 60 3.4 Docking studies of diazo and
    [Show full text]
  • Treatment of Cashew Extracts with Aspergillopepsin Reduces Ige Binding to Cashew Allergens
    Journal of Applied Biology & Biotechnology Vol. 4 (02), pp. 001-010, March-April, 2016 Available online at http://www.jabonline.in DOI: 10.7324/JABB.2016.40201 Treatment of cashew extracts with Aspergillopepsin reduces IgE binding to cashew allergens Cecily B. DeFreece1, Jeffrey W. Cary2, Casey C. Grimm2, Richard L. Wasserman3, Christopher P Mattison2* 1Department of Biology, Xavier University of Louisiana, New Orleans, LA, USA. 2USDA-ARS, Southern Regional Research Center, New Orleans, LA, USA. 3Allergy Partners of North Texas Research, Department of Pediatrics, Medical City Children’s Hospital, Dallas, Texas, USA. ARTICLE INFO ABSTRACT Article history: Enzymes from Aspergillus fungal species are used in many industrial and pharmaceutical applications. Received on: 21/10/2015 Aspergillus niger and Aspergillus oryzae were cultured on media containing cashew nut flour to identify secreted Revised on: 06/12/2015 proteins that may be useful as future food allergen processing enzymes. Mass-spectrometric analysis of secreted Accepted on: 20/12/2015 proteins and protein bands from SDS-PAGE gels indicated the presence of at least 63 proteins. The majority of Available online: 21/04/2016 these proteins were involved in carbohydrate metabolism, but there were also enzymes involved in lipid and protein metabolism. It is likely that some of these enzymes are specifically upregulated in response to cashew Key words: nut protein, and study of these enzymes could aid our understanding of cashew nut metabolism. Aspergillus, cashew, food Aspergillopepsin from A. niger was one of the proteolytic enzymes identified, and 6 distinct peptides were allergy, immunoglobulin E, matched to this protein providing 22% coverage of the protein.
    [Show full text]
  • Downloading the Nucleotide Sequences and Scanning Them Against the Database
    An in silico analysis, purification and partial kinetic characterisation of a serine protease from Gelidium pristoides A dissertation submitted in fulfilment of the requirement for the degree of Master of Science (MSc) Biochemistry by Zolani Ntsata Supervisor: Prof. Graeme Bradley 2020 Department of Biochemistry and Microbiology Declaration I, Zolani Ntsata (201106067), declare that this dissertation, entitled ‘An in silico analysis and kinetic characterisation of proteases from red algae’ submitted to the University of Fort Hare for the Master’s degree (Biochemistry) award, is my original work and has NOT been submitted to any other university. Signature: __________________ I, Zolani Ntsata (201106067), declare that I am highly cognisant of the University of Fort Hare policy on plagiarism and I have been careful to comply with these regulations. Furthermore, all the sources of information have been cited as indicated in the bibliography. Signature: __________________ I, Zolani Ntsata (201106067), declare that I am fully aware of the University of Fort Hare’s policy on research ethics, and I have taken every precaution to comply with these regulations. There was no need for ethical clearance. Signature: _________________ i Dedication I dedicate this work to my grandmother, Nyameka Mabi. ii Acknowledgements Above all things, I would like to give thanks to God for the opportunity to do this project and for the extraordinary strength to persevere in spite of the challenges that came along. I am thankful to my family, especially my grandmother, for her endless support. I would also like to acknowledge Prof Graeme Bradley for his supervision and guidance. Thanks to my friends and colleagues, especially Yanga Gogela and Ntombekhaya Nqumla, and the plant stress group for their help and support.
    [Show full text]
  • Investigating the Impact of Mpapr1, an Aspartic Protease from the Yeast Metschnikowia Pulcherrima, on Wine Properties
    THÈSE EN COTUTELLE PRÉSENTÉE POUR OBTENIR LE GRADE DE DOCTEUR DE L’UNIVERSITÉ DE BORDEAUX ET DE L’UNIVERSITÉ DE STELLENBOSCH ÉCOLE DOCTORALE DES SCIENCES DE LA VIE ET DE LA SANTÉ SPÉCIALITÉ ŒNOLOGIE FACULTY OF AGRISCIENCES Par Louwrens THERON ETUDE DE L’IMPACT DE MPAPR1, UNE PROTEASE ASPARTIQUE DE LA LEVURE METSCHNIKOWIA PULCHERRIMA, SUR LES PROPRIETES DU VIN Sous la direction de Benoit DIVOL et de Marina BELY Soutenue le 27 janvier 2017 Membres du jury: Mme. LE HENAFF-LE MARREC Claire, Professeur à l’université de Bordeaux Président M. MARANGON Matteo, Chargé de recherche à l’université de Padoue Rapporteur Mme. CAMARASA Carole, Chargée de recherche à l’INRA de Montpellier Rapporteur M. BAUER Florian, Professeur à l’université de Stellenbosch Examinateur Titre : Etude de l’impact de MpAPr1, une protéase aspartique de la levure Metschnikowia pulcherrima, sur les propriétés du vin Résumé : L'élimination des protéines est une étape clé lors de la production du vin blanc afin d'éviter l'apparition éventuelle d'un voile inoffensif mais inesthétique. Des solutions de rechange à l'utilisation de la bentonite sont activement recherchées en raison des problèmes technologiques, organoleptiques et de durabilité associés à son utilisation. Dans cette étude, MpAPr1, une protéase aspartique extracellulaire préalablement isolée et partiellement caractérisée à partir de la levure Metschnikowia pulcherrima, a été clonée et exprimée de manière hétérologue dans la levure Komagataella pastoris. Les propriétés enzymatiques de MpAPr1 ont été initialement caractérisées dans un extrait brut. Après plusieurs essais faisant appel à différentes techniques, MpAPr1 a été purifié avec succès par chromatographie échangeusede cations.
    [Show full text]
  • (A) Enzymes Derived from Animal Sources
    ANNEX (new entries are highlighted in yellow) PERMITTED ENZYMES (A) Enzymes derived from animal sources Enzyme EC Number Source Catalase 1.11.1.6 Bovine liver Lactoperoxidase 1.11.1.7 Bovine milk Bovine stomach; salivary glands or forestomach of calf, kid or lamb; porcine Lipase, triacylglycerol 3.1.1.3 or bovine pancreas Lysozyme 3.2.1.17 Egg whites Pancreatin (or pancreatic elastase) 3.4.21.36 Pancreas of the hog or ox Pepsin 3.4.23.1 Bovine or porcine stomach Phospholipase A2 3.1.1.4 Porcine pancreas Aqueous extracts from the fourth stomach of calves, kids, lambs, and adult Rennet 3.4.23.4 bovine animals, sheep and goats Thrombin 3.4.21.5 Bovine or porcine blood Trypsin 3.4.21.4 Porcine or bovine pancreas (B) Enzymes derived from plant sources Enzyme EC Number Source Alpha–amylase 3.2.1.1 Malted cereals Actinidin 3.4.22.14 Kiwifruit (Actinidia deliciosa) Malted cereals Beta-Amylase 3.2.1.2 Sweet potato (Ipomoea batatas) Bromelain 3.4.22.4 Pineapple fruit/stem (Ananas comosus and Ananas bracteatus (L)) Ficin 3.4.22.3 Ficus spp. Lipoxidase 1.13.11.12 Soyabean whey or meal Papain 3.4.22.2 Carica papaya (L) (Fam. Caricaceae) (C) Enzymes derived from microbial sources EC Enzyme Production organism Donor organism Donor gene Number 1,4-alpha-glucan branching 1,4-alpha-glucan branching 2.4.1.18 Bacillus subtilis Rhodothermus obamensis enzyme enzyme Alpha-acetolactate decarboxylase 4.1.1.5 Bacillus amyloliquefaciens Bacillus subtilis Bacillus subtilis Bacillus brevis Alpha-acetolactate decarboxylase Alpha-amylase 3.2.1.1 Aspergillus niger1 Aspergillus
    [Show full text]
  • Handbook of Proteolytic Enzymes Second Edition Volume 1 Aspartic and Metallo Peptidases
    Handbook of Proteolytic Enzymes Second Edition Volume 1 Aspartic and Metallo Peptidases Alan J. Barrett Neil D. Rawlings J. Fred Woessner Editor biographies xxi Contributors xxiii Preface xxxi Introduction ' Abbreviations xxxvii ASPARTIC PEPTIDASES Introduction 1 Aspartic peptidases and their clans 3 2 Catalytic pathway of aspartic peptidases 12 Clan AA Family Al 3 Pepsin A 19 4 Pepsin B 28 5 Chymosin 29 6 Cathepsin E 33 7 Gastricsin 38 8 Cathepsin D 43 9 Napsin A 52 10 Renin 54 11 Mouse submandibular renin 62 12 Memapsin 1 64 13 Memapsin 2 66 14 Plasmepsins 70 15 Plasmepsin II 73 16 Tick heme-binding aspartic proteinase 76 17 Phytepsin 77 18 Nepenthesin 85 19 Saccharopepsin 87 20 Neurosporapepsin 90 21 Acrocylindropepsin 9 1 22 Aspergillopepsin I 92 23 Penicillopepsin 99 24 Endothiapepsin 104 25 Rhizopuspepsin 108 26 Mucorpepsin 11 1 27 Polyporopepsin 113 28 Candidapepsin 115 29 Candiparapsin 120 30 Canditropsin 123 31 Syncephapepsin 125 32 Barrierpepsin 126 33 Yapsin 1 128 34 Yapsin 2 132 35 Yapsin A 133 36 Pregnancy-associated glycoproteins 135 37 Pepsin F 137 38 Rhodotorulapepsin 139 39 Cladosporopepsin 140 40 Pycnoporopepsin 141 Family A2 and others 41 Human immunodeficiency virus 1 retropepsin 144 42 Human immunodeficiency virus 2 retropepsin 154 43 Simian immunodeficiency virus retropepsin 158 44 Equine infectious anemia virus retropepsin 160 45 Rous sarcoma virus retropepsin and avian myeloblastosis virus retropepsin 163 46 Human T-cell leukemia virus type I (HTLV-I) retropepsin 166 47 Bovine leukemia virus retropepsin 169 48
    [Show full text]
  • Identification and Characterization of a Bacterial Glutamic Peptidase
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Copenhagen University Research Information System Identification and characterization of a bacterial glutamic peptidase Jensen, Kenneth; Østergaard, Peter R.; Wilting, Reinhard ; Lassen, Søren F. Published in: BMC Biochemistry DOI: 10.1186/1471-2091-11-47 Publication date: 2010 Document version Publisher's PDF, also known as Version of record Citation for published version (APA): Jensen, K., Østergaard, P. R., Wilting, R., & Lassen, S. F. (2010). Identification and characterization of a bacterial glutamic peptidase. BMC Biochemistry, 11(47). https://doi.org/10.1186/1471-2091-11-47 Download date: 07. Apr. 2020 Jensen et al. BMC Biochemistry 2010, 11:47 http://www.biomedcentral.com/1471-2091/11/47 RESEARCH ARTICLE Open Access Identification and characterization of a bacterial glutamic peptidase Kenneth Jensen1,2*, Peter R Østergaard1, Reinhard Wilting1, Søren F Lassen1 Abstract Background: Glutamic peptidases, from the MEROPS family G1, are a distinct group of peptidases characterized by a catalytic dyad consisting of a glutamate and a glutamine residue, optimal activity at acidic pH and insensitivity towards the microbial derived protease inhibitor, pepstatin. Previously, only glutamic peptidases derived from filamentous fungi have been characterized. Results: We report the first characterization of a bacterial glutamic peptidase (pepG1), derived from the thermoacidophilic bacteria Alicyclobacillus sp. DSM 15716. The amino acid sequence identity between pepG1 and known fungal glutamic peptidases is only 24-30% but homology modeling, the presence of the glutamate/ glutamine catalytic dyad and a number of highly conserved motifs strongly support the inclusion of pepG1 as a glutamic peptidase.
    [Show full text]
  • EUROPEAN COMMISSION Brussels, 28 April 2020 REGISTER of FOOD
    EUROPEAN COMMISSION DIRECTORATE-GENERAL FOR HEALTH AND FOOD SAFETY Food and feed safety, innovation Food processing technologies and novel foods Brussels, 28 April 2020 REGISTER OF FOOD ENZYMES TO BE CONSIDERED FOR INCLUSION IN THE UNION LIST Article 17 of Regulation (EC) No 1332/20081 provides for the establishment of a Register of all food enzymes to be considered for inclusion in the Union list. In accordance with that Article, the Register includes all applications which were submitted within the initial period fixed by that Regulation and which comply with the validity criteria laid down in accordance with Article 9(1) of (EC) No 1331/2008 establishing a common authorisation procedure for food additives, food enzymes and food flavourings2. The Register therefore lists all valid food enzyme applications submitted until 11 March 2015 except those withdrawn by the applicant before that date. Applications submitted after that date are not included in the Register but will be processed in accordance with the Common Authorisation Procedure. The entry of a food enzyme in the Register specifies the identification, the name, the source of the food enzyme as provided by the applicant and the EFSA question number under which the status of the Authority’s assessment can be followed3. As defined by Article 3 of Regulation (EC) No 1332/2008, ‘food enzyme’ subject to an entry in the Register, refers to a product that may contain more than one enzyme capable of catalysing a specific biochemical reaction. In the assessment process, such a food enzyme may be linked with several EFSA question numbers.
    [Show full text]