Chemistry: natural resins or derivatives; peptides or proteins; – Peptides of 3 to 100 amino acid residues – 25 or more amino acid residues in defined sequence
Reexamination Certificate
1995-06-01
2001-07-31
Prouty, Rebecca E. (Department: 1652)
Chemistry: natural resins or derivatives; peptides or proteins;
Peptides of 3 to 100 amino acid residues
25 or more amino acid residues in defined sequence
C536S023700, C530S825000, C435S849000
Reexamination Certificate
active
06268471
ABSTRACT:
TECHNICAL FIELD
This invention is related to the field of microbiology. Specifically, the invention relates to alteration of metabolic pathways in
Escherichia coli
and other bacteria.
BACKGROUND ART
Bacteria grown in the laboratory have three stages of growth, the lag phase in which little growth occurs, the log phase in which exponential growth occurs and the stationary phase in which growth ceases. During the transition into stationary phase, bacteria acquire numerous new physiological properties which enhance their ability to compete and survive under suboptimal conditions. For reviews, see Kolter (1992)
ASM News
58:75-79; Matin (1991)
Mol. Microbiol.
5:3-10; Matin et al. (1989)
Ann. Rev. Microbiol.
43:293-316; and Siegele and Kolter (1992)
J. Bacteriol.
174:345-348. In
Escherichia coli
the induction of several genes and operons in the stationary phase requires a putative sigma factor, katF or rpos. Bohannon et al. (1991)
J. Bacteriol.
173:4482-4492; Lange and Hengge-Aronis (1991)
Mol. Microbiol.
5:49-59; Matin (1991); and Schellhorn and Stones (1992)
J. Bacteriol.
174:4769-4776. The expression of stationary-phase genes such as mcbA for microcin production and glgCA for glycogen synthesis do not require katF. Bohannon et al. (1991).
Previous studies of factors that control the glycogen biosynthesis genes in
E. coli
showed that cyclic AMP (cAMP), cAMP receptor protein, and guanosine 3′-bisphosphate 5′-bisphosphate (ppGpp) stimulate the expression of the genes for the essential enzymes of the glycogen pathway, glgC (encoding ADPglucose pyrophosphorylase [EC 2.7.7.27]) and glgA (encoding glycogen synthase [EC 2.4.1.21]) which are apparently cotranscribed in an operon, glgCAY. Romeo et al. (1990)
Curr. Microbiol.
21:131-137; and Romeo and Priess (1989)
J. Bacteriol.
171:2773-2782. This operon also includes the gene encoding the catabolic enzyme glycogen phosphorylase [E.C 2.4.1.1], glgY or glgP. Romeo et al. (1988)
Gene
70:363-376; and Yu et al. (1988)
J. Biol. Chem.
263:13706-13711.
The gene glgB (encoding glycogen branching enzyme [EC 2.4.1.18]) is located upstream from glgCAY, apparently in an operon, glgBX, that includes a gene encoding a second catabolic enzyme. Baecker et al. (1986)
J. Biol. Chem.
261:8738-8743; and Romeo et al. (1988). Although the expression of the three biosynthetic genes is induced in stationary phase, glgB is transcribed independently of glgCA and is not regulated by cAMP-cAMP receptor protein or ppGpp. Preiss and Romeo (1989)
Adv. Microb. Physiol.
30:183-233; and Romeo and Preiss (1989). Four stationary-phase-induced transcripts have been mapped within the 0.5 kb upstream noncoding region of the glgc gene from
E. coli
, implying complex transcriptional regulation of glgCA. Romeo and Preiss (1989).
It would be highly advantageous to inhibit the metabolic pathway for glycogen biosynthesis and later reactions of gluconeogenesis in bacterial host cells, while stimulating anaplerotic reactions such as phosphoenolpyruvate carboxykinase in order to channel more carbon atoms into desired products such as aromatic amino acids.
DISCLOSURE OF THE INVENTION
The invention is directed to the the csrA gene, the protein encoded thereby and methods of use thereof. Modulation of csrA gene expression is useful in regulating expression of downstream metabolic products and alone, or in conjunction with other mutations, is suitable for use in production of bacterial products.
REFERENCES:
patent: WO 91/17247 (1991-11-01), None
White et al., “Phylogenetic disitribution of the regulatory gene csrA among eubacteria”Gene(1996) 182:221-223.
Romeo et al., “Carbon storage regulator; CSRA or ZFIA;Escherichia coli” Swissport Sequence Database(Jul. 1, 1993) Heidelberg, BRD, XP002040654, Accession No. P31803.
Romeo et al., “Escherichia colicarbon storage regulator (csrA) gene, complete cds; alaS gene, 3′ end, andserv promoter regulator”EMBL Sequence Database(Jun. 15, 1993) Heidelberg, BRD, XP002040655, Accession No. L07596.
Blatch et al., “Nucleotide sequence and analysis of theVibrio alginolyticussucrose uptake-encoding region”Gene(1990) 95:17-23.
Cui et al., “Identification of a global repressor gene, rsmA, ofErwinia carotovorasubsp.carotovorathat controls extracellular enzymes, N-(3-oxohexanoyl)-L-homoserine lactone, and pathogenicity in soft-rotting Erwinia spp.”J. Bacteriol.(1995) 177(17):5108-5115.
Liu et al., “The global regulator CsrA ofEscherichia coliis a specific mRNA-binding protein”J. Bacteriol.(1997) 179(14):4639-4642.
Liu et al., “The product of the pleiotropicEscherichia coligene csrA modulates glycogen biosynthesis via effects on mRNA stability”J. Bacteriol.(1995) 177(10):2663-2672.
Liu et al., “The RNA molecule CsrB binds to the global regulatory protein CsrA and antagonizes its activity inEscherichia coli” J. Biol. Chem.(1997) 272(28):17502-17510.
Mukherjee et al., “Global regulation in Erwinia species byErwinia carotovorarsmA, a homologue ofEscherichia colicsrA: repression of secondary metabolites, pathogenicity and hypertensive reaction”Microbiol.(1996) 142:427-434.
Murayama et al., “Evidence for involvement ofEscherichia coligenes pmbA, csrA and a previously unrecognized gene tldD, in the control of DNA gyrase by letD (ccdB) of sex factor F”J. Mol. Biol.(1996) 256:483-502.
Romeo, “Post-transcriptional regulation of bacterial carbohydrate metabolism: evidence that the gene product CsrA is a global mRNA decay factor”Res. Microbiol.(1996) 147(6-7):505-512.
Sabnis et al., “Pleiotropic regulation of central carbohydrate metabolism inEscherichia colivia the gene csrA”J. Biol. Chem.(1995) 270:29096-29104.
Yang et al., “Coordinate genetic regulation of glycogen catabolism and biosynthesis inEscherichia colivia the CsrA gene product”J. Bacteriol.(1996) 178:000-000 (prepublication copy: pp. a-f).
Morrison & Foerster / LLP
Prouty Rebecca E.
University of North Texas Health Science Center at Forth Worth
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