Dyadic International (usa), Inc.

Nederlandse Organisatie Voor Toegepast-natuurwetenschappelijk Onderzoek Tno

Dutch Dna Biotech B. V.

Novozymes A/s

Danisco US Inc.

Roal Oy

Dyadic International, Inc.

Other

Institut Francais Du Pe'trole

Institut National De La Recherche Agronomique

Dyadic Nederland B.v.

Peter Jan Punt

Mark Aaron Emalfarb

Richard Paul Burlingame

Arkady Panteleimonovich Sinitsyn

Jan Cornelis Verdoes

Christine Marie Pynnonen

Maria Johanna Van Der Werf

Philip Terry Olson

Martine Parriche

Cornelia Van Zeijl

Cornelius Van Den Hondel

Jean Christophe Bousson

Cornelia Marie Johanna Van Zeijl

Elena Vlasenko

Jacob B Visser

Marja T Paloheimo

Jari Vehmaanperä

Terhi Puranen

Kari Juntunen

Tove Christensen

Carsten Mailand Hjort

Richard P Burlingame

Alexander Vasilievich Gusakov

Susanna Mäkinen

Jan Wery

Johannes Heinrich Visser

Cornelia Maria Johanna Van Zeijl

Sandra Wihelmina Agnes Hinz

Mark Gosink

Zhijie Jiang

Phillip T Olson

Frank H J Schuren

Cornelia Maria Van Zeijl

Marcel Asther

Frédéric Monot

Eric Record

Jean-Pierre Belaïch

Nicole Van Luijk

Anthony Levasseur

David Navarro

Jacoba Van Der Meij, Legal Representative

An Li

Cees Van Den Hondel

Mark Aaron Emalfrab

Martinus Petrus Maria Caspers

Jacoba Van Der Meij

Cees A M J J Van Den Hondel

Susanna MÄKINEN

Jari VehmaanperÄ

The present description is related to the field of protein production. It introduces novel host cells with low protease activity, a novel protease regulator, its use in expression systems and protein production, and a method of producing host cells for protein production.

Methods for controlling protease production

The present invention provides a new fungal production system comprising a fungal host strain ofwherein the endogenous cellulase secretion is less than 20% of the endogenous cellulase secretion ofstrain UV 18-25. Preferably, also the secretion of endogenous protease, endogenous &#x3b2;-glucanase and endogenous cellobiohydrolase is less than 20% of the secretion ofstrain UV 18-25. Furthermore, fungal host strains are provided wherein several genes have been disrupted. According to another aspect of the invention a method for homologous and/or heterologous production of a pure protein with a purity of higher than 75%, comprising expressing a gene encoding said protein in the strains according to the invention have been described. Furthermore, a method for production of artificial protein mixes comprising expressing a gene encoding each of said proteins in a strain according to the invention have been disclosed. Finally a method for simplified screening of strains functionally expressing a desired enzyme by application of said strains have been provided.

Fungal production system

The invention relates to the use of a cytosolic citric acid synthase for the heterologous production of citrate outside the mitochondrion of a micro-organism or algae, wherein the protein is selected fromAn08g10920, An01g09940, An09g03570, or an ortholog of these genes. Such production is achieved by introducing the nucleic acid encoding such a protein into a suitable host cell. Preferably the protein isAn08g10920, An01g09940, An09g03570 or an ortholog thereof, more particularly, wherein such an ortholog is chosen from the group of proteins listed in FIG.and proteins having a percentage identity of 70%, more preferably 75%, more preferably 80%, more preferably 85%, more preferably 90%, more preferably 95%, more preferably 98%, more preferably 99% with An08g10920, An01g09940 or An09g03570.

Organic acid pathway

The invention is generally directed to modified filamentous fungal host cells comprising one or more nucleic acids encoding one or more polypeptides under the control of one or more promoters that are functional in said cells. Methods of using the modified cells to express one or more polypeptides are also disclosed, including methods of screening cells transformed with one or more expression vectors comprising nucleic acids derived from synthetic or genomic nucleic acids including, cDNAs. Methods of purifying one or more polypeptides or complexes comprising one or more polypeptides expressed in the modified cells, intended for use as substrates in structure/function studies, as therapeutic agents, as diagnostic reagents, or as human or animal vaccines, are also disclosed.

Expression and high-throughput screening of complex expressed DNA libraries in filamentous fungi

The invention relates to a nucleic acid sequence encoding an itaconate transporting Major Facilitator Superfamily Transporter (MFST) gene sequence and the protein encoded thereby. Preferably said sequence is the nucleic acid that comprises the sequence of ATEG_09972.1 ofor homologues thereof. Overexpression of the protein enhances the production of itaconic acid in micro-organisms.

Production of itaconic acid

The invention relates to a nucleic acid sequence encoding anmitochondrial tricarboxylic acid transporter that can be used in the production of itaconic acid in micro-organisms. Preferably said transporter protein is the protein encoded by the nucleic acid which is located on a chromosome segment ofthat also harbors other genes involved in the itaconic acid biosynthesis and the lovastatin biosynthesis. Also, vectors, hosts and transformed micro-organisms are part of the invention.

Methods to digest carbohydrates, especially lignocelluloses and hemicelluloses, using fungal proteins previously not recognized as having this activity are described.

Cellulose and ligno-cellulose active proteins

A novel transformation system in the field of filamentous fungal hosts for expressing and secreting heterologous proteins or polypeptides is described. The invention also covers a process for producing large amounts of polypeptide or protein in an economical manner. The system comprises a transformed or transfected fungal strain of the genus, more particularly ofand mutants or derivatives thereof. It also covers transformants containingcoding sequences, as well expression-regulating sequences ofgenes. Also provided are novel fungal enzymes and their encoding sequences and expression-regulating sequences.

Transformation system in the field of filamentous fungal hosts

The invention provides a method for the expression and subsequent screening of DNA libraries, particularly synthetic, genomic, and cDNA libraries, in filamentous fungal hosts. In particular, the invention provides vectors, host strains, and a method for the expression and screening of complex DNA libraries, including, but not limited to, combinatory (combinatorial) libraries expressing one, two or more variable constituents and/or prepared from two or more sublibraries (e.g., for the expression and screening of immunoglobulin (including fragments and derivatives of whole immunoglobulin proteins) and other receptor or complex DNA libraries or libraries of libraries). The invention is useful for the expression and screening for a large variety of proteins and protein complexes, including human proteins. The present invention also relates to novel fungal protease sequences.

The invention relates to a nucleic acid sequence encoding an itaconate transporting Major Facilitator Superfamily Transporter (MFST) gene sequence and the protein encoded thereby. Preferably said sequence is the nucleic acid that comprises the sequence of ATEG_09972.1 ofor homologues thereof. Overexpression of the protein enhances the production of itaconic acid in microorganisms.

This invention relates to novel enzymes and novel methods for producing the same. More specifically this invention relates to a variety of fungal enzymes. Nucleic acid molecules encoding such enzymes, compositions, recombinant and genetically modified host cells, and methods of use are described. The invention also relates to a method to convert lignocellulosic biomass to fermentable sugars with enzymes that degrade the lignocellulosic material and novel combinations of enzymes, including those that provide a synergistic release of sugars from plant biomass. The invention also relates to a method to release cellular content by degradation of cell walls. The invention also relates to methods to use the novel enzymes and compositions of such enzymes in a variety of other processes, including washing of clothing, detergent processes, biorefining, deinking and biobleaching of paper and pulp, and treatment of waste streams.

BX11 enzymes having xylosidase activity

Growing community of inventors

Houten, Netherlands

Peter Jan Punt