JBC Ideal method for primary cell transfection

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

This Article
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Shinabarger, D.
Right arrow Articles by Chakrabarty, A. M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Shinabarger, D.
Right arrow Articles by Chakrabarty, A. M.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

J. Biol. Chem., Vol. 266, Issue 4, 2080-2088, Feb, 1991

Purification and characterization of phosphomannose isomerase-guanosine diphospho-D-mannose pyrophosphorylase. A bifunctional enzyme in the alginate biosynthetic pathway of Pseudomonas aeruginosa

D Shinabarger, A Berry, TB May, R Rothmel, A Fialho and AM Chakrabarty
Department of Microbiology and Immunology, University of Illinois College of Medicine, Chicago 60612.

We report here the purification and characterization of phosphomannose isomerase-guanosine 5'-diphospho-D-mannose pyrophosphorylase, a bifunctional enzyme (PMI-GMP) which catalyzes both the phosphomannose isomerase (PMI) and guanosine 5'-diphospho-D-mannose pyrophosphorylase (GMP) reactions of the Pseudomonas aeruginosa alginate biosynthetic pathway. The PMI and GMP activities co-eluted in the same protein peak through successive fractionation on hydrophobic interaction, ion exchange, and gel filtration chromatography. The purified enzyme migrated as a 56,000 molecular weight protein on sodium dodecyl sulfate- polyacrylamide gel electrophoresis, and the native protein migrated as a monomer of 54,000 molecular weight upon gel filtration chromatography. The apparent Km for D-mannose 6-phosphate was 3.03 mM, and the Vmax was 830 nmol/min/mg of enzyme. For the GMP forward reaction, apparent Km values of 20.5 microM and 29.5 microM for D- mannose 1-phosphate and GTP, respectively, were obtained from double reciprocal plots. The GMP forward reaction Vmax (5,680 nmol/min/mg of enzyme) was comparable to the reverse reaction Vmax (5,170 nmol/min/mg of enzyme), and the apparent Km for GDP-D-mannose was determined to be 14.2 microM. Both reactions required Mg2+ activation, but the PMI reaction rate was 4-fold higher with Co2+ as the activator. PMI (but not GMP) activity was sensitive to dithiothreitol, indicating the involvement of disulfide bonds to form a protein structure capable of PMI activity. DNA sequencing of a cloned mutant algA gene from P. aeruginosa revealed that a point mutation at nucleotide 961 greatly decreased the levels of both PMI and GMP in a crude extract.
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 MicrobiologyHome page
L. L. Oglesby, S. Jain, and D. E. Ohman
Membrane topology and roles of Pseudomonas aeruginosa Alg8 and Alg44 in alginate polymerization
Microbiology, June 1, 2008; 154(6): 1605 - 1615.
[Abstract] [Full Text] [PDF]

Home page
 J. Bacteriol.Home page
M. Gimmestad, H. Sletta, H. Ertesvag, K. Bakkevig, S. Jain, S.-j. Suh, G. Skjak-Braek, T. E. Ellingsen, D. E. Ohman, and S. Valla
The Pseudomonas fluorescens AlgG Protein, but Not Its Mannuronan C-5-Epimerase Activity, Is Needed for Alginate Polymer Formation
J. Bacteriol., June 15, 2003; 185(12): 3515 - 3523.
[Abstract] [Full Text] [PDF]

Home page
 Microbiol. Mol. Biol. Rev.Home page
M. A. Ballicora, A. A. Iglesias, and J. Preiss
ADP-Glucose Pyrophosphorylase, a Regulatory Enzyme for Bacterial Glycogen Synthesis
Microbiol. Mol. Biol. Rev., June 1, 2003; 67(2): 213 - 225.
[Abstract] [Full Text] [PDF]

Home page
 MicrobiologyHome page
R. C. Keith, L. M. W. Keith, G. Hernandez-Guzman, S. R. Uppalapati, and C. L. Bender
Alginate gene expression by Pseudomonas syringae pv. tomato DC3000 in host and non-host plants
Microbiology, May 1, 2003; 149(5): 1127 - 1138.
[Abstract] [Full Text] [PDF]

Home page
 J. Bacteriol.Home page
M. J. Franklin and D. E. Ohman
Mutant Analysis and Cellular Localization of the AlgI, AlgJ, and AlgF Proteins Required for O Acetylation of Alginate in Pseudomonas aeruginosa
J. Bacteriol., June 1, 2002; 184(11): 3000 - 3007.
[Abstract] [Full Text] [PDF]

Home page
 Microbiol. Mol. Biol. Rev.Home page
J. M. Dunwell, S. Khuri, and P. J. Gane
Microbial Relatives of the Seed Storage Proteins of Higher Plants: Conservation of Structure and Diversification of Function during Evolution of the Cupin Superfamily
Microbiol. Mol. Biol. Rev., March 1, 2000; 64(1): 153 - 179.
[Abstract] [Full Text] [PDF]

Home page
 Microbiol. Mol. Biol. Rev.Home page
H. L. Rocchetta, L. L. Burrows, and J. S. Lam
Genetics of O-Antigen Biosynthesis in Pseudomonas aeruginosa
Microbiol. Mol. Biol. Rev., September 1, 1999; 63(3): 523 - 553.
[Abstract] [Full Text] [PDF]

Home page
 J. Bacteriol.Home page
M. K. Fakhr, A. Peñaloza-Vázquez, A. M. Chakrabarty, and C. L. Bender
Regulation of Alginate Biosynthesis in Pseudomonas syringae pv. syringae
J. Bacteriol., June 1, 1999; 181(11): 3478 - 3485.
[Abstract] [Full Text]

Home page
 Proc. Natl. Acad. Sci. USAHome page
P. L. Conklin, S. R. Norris, G. L. Wheeler, E. H. Williams, N. Smirnoff, and R. L. Last
Genetic evidence for the role of GDP-mannose in plant ascorbic acid (vitamin C) biosynthesis
PNAS, March 30, 1999; 96(7): 4198 - 4203.
[Abstract] [Full Text] [PDF]


HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
 All ASBMB Journals   Molecular and Cellular Proteomics 
 Journal of Lipid Research   ASBMB Today 
Copyright © 1991 by the American Society for Biochemistry and Molecular Biology.