Iogen Energy Corporation

Codexis, Inc.

Brian R Scott

John J Tomashek

James A Lavigne

Chengsong Liu

Christopher Hill

Andrew Shaw

Ish Kumar Dhawan

Dipnath Baidyaroy

Theresa C White

Patrick St-Pierre

Nabil Masri

Oleh Tanchak

Amala Chokshi

Daniel Kolczynski

Martine Whissel

Provided are modified cellulase enzymes exhibiting increase cellulose-hydrolyzing activity in the presence of lignin and/or reduced binding to lignin comprising modified linker peptides comprising one or more amino acid substitutions, insertions, or deletions that result in (a) a decrease in the calculated isoelectric point of the linker peptide and/or (b) an increase in the ratio of threonine:serine in the linker peptide relative to a parental linker peptide from which said modified linker peptide is derived. Also provided are genetic constructs comprising nucleic acid sequences encoding for modified cellulase enzymes, methods for the production of the modified cellulase enzymes from host strains and a process for hydrolysing cellulose with the modified cellulases in the presence of lignin.

Cellulase enzymes having a modified linker and reduced lignin binding

Provided is a modified Family 1 carbohydrate binding module (CBM) comprising amino acid substitutions at one or more of positions 10, 11, 12, 14, 17, 21, 24, 29, 31, 33, and 37, said position determined from alignment of a Family 1 CBM amino acid sequence with SEQ ID NO: 30, and exhibiting from about 50% to about 99.9% amino acid sequence identity to SEQ ID NO: 30. Also provided are modified glycosidase enzymes comprising the modified Family 1 CBM, genetic constructs and genetically modified microbes for expressing the modified Family 1 CBM or modified glycosidase enzyme. The modified Family 1 CBM confers reduced lignin binding and/or increased hydrolyzing activity in the presence of lignin to the modified glycosidase enzyme, which may be used in a process for hydrolyzing cellulose or hemicellulose in the presence of lignin.

Carbohydrate binding modules with reduced binding to lignin

Provided are modified cellulase enzymes exhibiting increase cellulose-hydrolyzing activity in the presence of lignin and/or reduced binding to lignin comprising modified linker peptides comprising one or more amino acid substitutions, insertions, or deletions that result in (a) a decrease in the calculated isoelectric point of the linker peptide and/or (b) an increase in the ratio of threonine:serine in the linker peptide relative to a parental linker peptide from which said modified linker peptide is derived. Also provided are genetic constructs comprising nucleic acid sequences encoding for modified cellulase enzymes, methods for the production of the modified cellulase enzymes from host strains and a process for hydrolyzing cellulose with the modified cellulases in the presence of lignin.

Carbohydate binding modules with reduced binding to lignin

Provided are modified beta-glucosidase enzymes, derived from theCel3A beta-glucosidase, that exhibit improvements in one or more kinetic parameters (K, K, k) comprising amino acid substitutions at one or more of positions 43, 101, 260 and 543. Also provided are genetic constructs comprising nucleotide sequences encoding for modified beta-glucosidase enzymes, methods for the production of modified beta-glucosidase enzymes from host strains and the use of the modified beta-glucosidase enzymes in the hydrolysis of cellulose.

Beta-glucosidase enzymes

The present application provides genetically modified fungal organisms that produce enzyme mixtures exhibiting enhanced hydrolysis of cellulosic material to glucose, enzyme mixtures produced by the genetically modified fungal organisms, and processes for producing glucose from cellulose using such enzyme mixtures.

Compositions and methods for production of fermentable sugars

A modifiedFamily 6 (TrCel6A) cellulase enzyme comprising amino acid substitutions at one or more positions selected from the group consisting of 129, 322, 363 and 410 of SEQ ID NO: 1 is provided. Genetic constructs and genetically modified microbes comprising nucleic sequences encoding the modified TrCel6a cellulase are also provided. The modified TrCel6A cellulase of the invention display at least a 15% decrease in inactivation by lignin relative to a parental TrCel6A cellulase from which the modified TrCel6A is derived. Such cellulases find use in a variety of applications in industry requiring enzymatic hydrolysis of cellulose in the presence of lignin, e.g., the hydrolysis of pretreated lignocellulosic feedstocks for the production of fermentable sugars, sugar alcohols and fuel alcohols.

Family 6 cellulase with decreased inactivation by lignin

Provided is an enzyme mixture for hydrolyzing a pretreated lignocellulosic feedstock to soluble sugars. The enzyme mixture comprises EG4 at a fractional concentration (f) of about 0.25 to about 0.83 (w/w), Swollenin at a fractional concentration (f) of about 0 to about 0.66 (w/w), and Cip1 at a fractional concentration (f) of 0 to about 0.33 measured relative to all accessory enzymes present in the enzyme mixture. Also provided are processes for converting a pretreated lignocellulosic feedstock to soluble sugars using the enzyme mixtures, and methods of using and producing such enzyme mixtures.

Growing community of inventors

Richmond, CA, United States of America

Brian R Scott