Directed evolution of metabolically engineered Escherichia coli for carotenoid production

C. Wang, Min-Kyu Oh, J. C. Liao

Research output: Contribution to journalArticle

102 Citations (Scopus)

Abstract

We have previously introduced a reconstructed isoprenoid pathway into Escherichia coli that exhibits amplified biosynthetic flux to geranylgeranyl diphosphate (GGPP), a common isoprenoid precursor. It was shown that GGPP synthase is an important rate-controlling enzyme in this reconstructed isoprenoid pathway. In this investigation, we applied directed evolution to GGPP synthase from Archaeoglobus fulgidus to enable the enhanced production of carotenoids in metabolically engineered E. coli. Eight mutants were isolated, and the best one increased lycopene production by 100%. Among the mutants that were isolated, mutation points were clustered in four "hot regions". The "hottest" region is located in the sequence upstream of the coding region, which presumably improves the expression level of the enzyme. The other three are within the coding sequence and are believed to improve the enzyme-specific activity in E coli. These results demonstrate that modulating both enzymatic expression and specific activity are important for optimizing the metabolic flux distribution.

Original languageEnglish
Pages (from-to)922-926
Number of pages5
JournalBiotechnology Progress
Volume16
Issue number6
DOIs
Publication statusPublished - 2000 Dec 1
Externally publishedYes

Fingerprint

isoprenoids
Terpenes
Carotenoids
Farnesyltranstransferase
carotenoids
Escherichia coli
Archaeoglobus
Archaeoglobus fulgidus
Enzymes
enzymes
mutants
point mutation
lycopene
Point Mutation
directed evolution
geranylgeranyl diphosphate synthase

ASJC Scopus subject areas

  • Food Science
  • Biotechnology
  • Microbiology

Cite this

Directed evolution of metabolically engineered Escherichia coli for carotenoid production. / Wang, C.; Oh, Min-Kyu; Liao, J. C.

In: Biotechnology Progress, Vol. 16, No. 6, 01.12.2000, p. 922-926.

Research output: Contribution to journalArticle

@article{252506ba9ce24a54ab2ccdc99d6ca740,
title = "Directed evolution of metabolically engineered Escherichia coli for carotenoid production",
abstract = "We have previously introduced a reconstructed isoprenoid pathway into Escherichia coli that exhibits amplified biosynthetic flux to geranylgeranyl diphosphate (GGPP), a common isoprenoid precursor. It was shown that GGPP synthase is an important rate-controlling enzyme in this reconstructed isoprenoid pathway. In this investigation, we applied directed evolution to GGPP synthase from Archaeoglobus fulgidus to enable the enhanced production of carotenoids in metabolically engineered E. coli. Eight mutants were isolated, and the best one increased lycopene production by 100{\%}. Among the mutants that were isolated, mutation points were clustered in four {"}hot regions{"}. The {"}hottest{"} region is located in the sequence upstream of the coding region, which presumably improves the expression level of the enzyme. The other three are within the coding sequence and are believed to improve the enzyme-specific activity in E coli. These results demonstrate that modulating both enzymatic expression and specific activity are important for optimizing the metabolic flux distribution.",
author = "C. Wang and Min-Kyu Oh and Liao, {J. C.}",
year = "2000",
month = "12",
day = "1",
doi = "10.1021/bp000124f",
language = "English",
volume = "16",
pages = "922--926",
journal = "Biotechnology Progress",
issn = "8756-7938",
publisher = "John Wiley and Sons Ltd",
number = "6",

}

TY - JOUR

T1 - Directed evolution of metabolically engineered Escherichia coli for carotenoid production

AU - Wang, C.

AU - Oh, Min-Kyu

AU - Liao, J. C.

PY - 2000/12/1

Y1 - 2000/12/1

N2 - We have previously introduced a reconstructed isoprenoid pathway into Escherichia coli that exhibits amplified biosynthetic flux to geranylgeranyl diphosphate (GGPP), a common isoprenoid precursor. It was shown that GGPP synthase is an important rate-controlling enzyme in this reconstructed isoprenoid pathway. In this investigation, we applied directed evolution to GGPP synthase from Archaeoglobus fulgidus to enable the enhanced production of carotenoids in metabolically engineered E. coli. Eight mutants were isolated, and the best one increased lycopene production by 100%. Among the mutants that were isolated, mutation points were clustered in four "hot regions". The "hottest" region is located in the sequence upstream of the coding region, which presumably improves the expression level of the enzyme. The other three are within the coding sequence and are believed to improve the enzyme-specific activity in E coli. These results demonstrate that modulating both enzymatic expression and specific activity are important for optimizing the metabolic flux distribution.

AB - We have previously introduced a reconstructed isoprenoid pathway into Escherichia coli that exhibits amplified biosynthetic flux to geranylgeranyl diphosphate (GGPP), a common isoprenoid precursor. It was shown that GGPP synthase is an important rate-controlling enzyme in this reconstructed isoprenoid pathway. In this investigation, we applied directed evolution to GGPP synthase from Archaeoglobus fulgidus to enable the enhanced production of carotenoids in metabolically engineered E. coli. Eight mutants were isolated, and the best one increased lycopene production by 100%. Among the mutants that were isolated, mutation points were clustered in four "hot regions". The "hottest" region is located in the sequence upstream of the coding region, which presumably improves the expression level of the enzyme. The other three are within the coding sequence and are believed to improve the enzyme-specific activity in E coli. These results demonstrate that modulating both enzymatic expression and specific activity are important for optimizing the metabolic flux distribution.

UR - http://www.scopus.com/inward/record.url?scp=0034518317&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0034518317&partnerID=8YFLogxK

U2 - 10.1021/bp000124f

DO - 10.1021/bp000124f

M3 - Article

C2 - 11101317

AN - SCOPUS:0034518317

VL - 16

SP - 922

EP - 926

JO - Biotechnology Progress

JF - Biotechnology Progress

SN - 8756-7938

IS - 6

ER -