Butamax Advanced Biofuels LLC

E.i. Dupont De Nemours and Company

Butamax (Tm) Advanced Biofuels LLC

Jean-Francois Tomb

Michael Dauner

Qiong Cheng

Vasantha Nagarajan

Lori Ann Maggio-Hall

Perry G Caimi

Andrew C Eliot

Mark J Nelson

Charles Edwin Nakamura

Gail K Donaldson

Steven Cary Rothman

Stephen K Picataggio

Michael G Bramucci

Gopal K Chotani

Rick Weizhang Ye

Pierre E Rouviere

Caroline M Peres

Luan Tao

Dennis Flint

Daniel P O'Keefe

Paul Veikko Viitanen

Larry Cameron Anthony

James M Odom

Siqun Wang

Mario W Chen

Wonchul Suh

Mattheos Koffas

Jahnavi Chandra Prasad

Kelley C Norton

Kristy N Kostichka

Yuying Zhang

Ritu Bhalla

Andreas Schenzle

Yougen Li

Tina K Van Dyk

Der-Ing Liao

Zheng You

Kristin Ruebling-Jass

Daniel P Okeefe

Laura McCole

Vasantha Nagarajan Nigarajan

The present invention is related to recombinant host cells comprising: (i) at least one deletion, mutation, and/or substitution in an endogenous gene encoding a polypeptide that converts pyruvate to acetaldehyde, acetyl-phosphate or acetyl-CoA; and (ii) a heterologous polynucleotide encoding a polypeptide having phosphoketolase activity. The present invention is also related to recombinant host cells further comprising (iii) a heterologous polynucleotide encoding a polypeptide having phosphotransacetylase activity.

Recombinant host cells comprising phosphoketalase

Provided herein are glycerol-3-phosphate dehydrogenase (GPD) enzymes with increased Kfor NADH and GPD enzymes with substantially the same affinity for NADH and NADPH and/or are feedback inhibited by glycerol-3-phosphate. Also provided herein are recombinant microorganisms comprising a heterologous gene encoding GPD and a deletion or disruption in an endogenous gene encoding GPD. Also provided are recombinant microorganisms comprising a heterologous gene encoding GPD and a butanol biosynthetic pathway. Further provided are methods of producing butanol comprising providing the recombinant microorganisms described herein and contacting the recombinant microorganism with at least one fermentable carbon substrate under conditions wherein butanol is produced.

Glycerol 3-phosphate dehydrogenase for butanol production

The present invention is related to recombinant host cells comprising: (i) at least one deletion, mutation, and/or substitution in an endogenous gene encoding a polypeptide that converts pyruvate to acetaldehyde, acetyl-phosphate, or acetyl-CoA; and (ii) a heterologous polynucleotide encoding a polypeptide having phosphoketolase activity. The present invention is also related to recombinant host cells further comprising (iii) a heterologous polynucleotide encoding a polypeptide having phosphotransacetylase activity.

Recombinant host cells comprising phosphoketolases

Glycerol 3- phosphate dehydrogenase for butanol production

Yeast strains were engineered that have increased activity of heterologous proteins that require binding of an Fe&#x2014;S cluster for their activity. The yeast strains have reduced activity of an endogenous Fe&#x2014;S protein. Activities of heterologous fungal or plant 2Fe-2S dihydroxy-acid dehydratases and Fe&#x2014;S propanediol dehydratase reactivase were increased for increased production of products made using biosynthetic pathways including these enzymes, such as valine, isoleucine, leucine, pantothenic acid (vitamin B5), isobutanol, 2-butanone and 2-butanol.

Recombinant yeast host cells comprising FE-S cluster proteins

Recombinant bacteria capable of metabolizing sucrose are described. The recombinant bacteria comprise in their genome or on at least one recombinant construct, a novel nucleotide sequence encoding a polypeptide having sucrose transporter activity and a nucleotide sequence encoding a polypeptide having sucrose hydrolase activity. These nucleotide sequences are each operably linked to the same or a different promoter. Recombinant bacteria capable of metabolizing sucrose to produce glycerol and/or glycerol-derived products such as 1,3-propanediol and 3-hydroxypropionic acid are also described.

Recombinant bacteria comprising novel sucrose transporters

Methods for the fermentive production of four carbon alcohols are provided. Specifically, butanol, preferably 2-butanol is produced by the fermentive growth of a recombinant bacteria expressing a 2-butanol biosynthetic pathway. The recombinant microorganisms and methods of the invention can also be adapted to produce 2-butanone, an intermediate in the 2-butanol biosynthetic pathways disclosed herein.

Fermentive production of four carbon alcohols

Recombinant host cell comprising a diol dehydratase

A group of bacterial dihydroxy-acid dehydratases having a [2Fe-2S] cluster was discovered. Bacterial [2Fe-2S] DHADs were expressed as heterologous proteins in bacteria and yeast cells, providing DHAD activity for conversion of 2,3-dihydroxyisovalerate to &#x3b1;-ketoisovalerate or 2,3-dihydroxymethylvalerate to &#x3b1;-ketomethylvalerate. Isobutanol and other compounds may be synthesized in pathways that include bacterial [2Fe-2S] DHAD activity.

Identification and use of bacterial [2Fe-2S] dihydroxy-acid dehydratases

Methods for the fermentive production of four carbon alcohols are provided. Specifically, butanol, preferably 2-butanol is produced by the fermentive growth of a recombinant bacteria expressing a 2-butanol biosynthetic pathway. The recombinant microorganisms and methods of the invention can also be adapted to produce 2-butanone, an intermediate in the 2-butanol biosynthetic pathways disclosed herein. Specifically disclosed herein are the use of coenzyme B-independent butanediol dehydratases that catalyzes the substrate to product conversion of 2,3-butanediol to 2-butanone in the process of producing 2-butanol and 2-butanone.

is unable to synthesize pantothenic acid and requires this essential vitamin in growth medium.strains transformed with an operon for expression of 2-dehydropantoate reductase and aspartate 1-decarboxylase were able to grow in medium lacking pantothenic acid. These strains may be used for ethanol production without pantothenic acid supplementation in seed culture and fermentation media.

Pantothenic acid biosynthesis in zymomonas

Xylose-utilizingstrains studied were found to accumulate ribulose when grown in xylose-containing media. Engineering these strains to increase ribose-5-phosphate isomerase activity led to reduced ribulose accumulation, improved growth, improved xylose utilization, and increased ethanol production.

Xylose utilization in recombinant zymomonas

Recombinant yeast host cell with Fe-S cluster proteins and methods of using thereof

Methods for the evolution of NADPH binding ketol-acid reductoisomerase enzymes to acquire NADH binding functionality are provided. Specific mutant ketol-acid reductoisomerase enzymes isolated fromthat have undergone co-factor switching to bind NADH are described.

Ketol-acid reductoisomerase using NADH

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Wilmington, DE, United States of America

Jean-Francois Tomb