Unraveling the Leloir Pathway of Bifidobacterium Bifidum: Significance of the Uridylyltransferases

Overview

Journal Appl Environ Microbiol

Specialties Environmental Health
Microbiology

Date 2013 Sep 10

PMID 24014529

Citations 10

Authors

Frederik De Bruyn

Joeri Beauprez

Jo Maertens

Wim Soetaert

Marjan De Mey

Affiliations

Soon will be listed here.

Abstract

The GNB/LNB (galacto-N-biose/lacto-N-biose) pathway plays a crucial role in bifidobacteria during growth on human milk or mucin from epithelial cells. It is thought to be the major route for galactose utilization in Bifidobacterium longum as it is an energy-saving variant of the Leloir pathway. Both pathways are present in B. bifidum, and galactose 1-phosphate (gal1P) is considered to play a key role. Due to its toxic nature, gal1P is further converted into its activated UDP-sugar through the action of poorly characterized uridylyltransferases. In this study, three uridylyltransferases (galT1, galT2, and ugpA) from Bifidobacterium bifidum were cloned in an Escherichia coli mutant and screened for activity on the key intermediate gal1P. GalT1 and GalT2 showed UDP-glucose-hexose-1-phosphate uridylyltransferase activity (EC 2.7.7.12), whereas UgpA showed promiscuous UTP-hexose-1-phosphate uridylyltransferase activity (EC 2.7.7.10). The activity of UgpA toward glucose 1-phosphate was about 33-fold higher than that toward gal1P. GalT1, as part of the bifidobacterial Leloir pathway, was about 357-fold more active than GalT2, the functional analog in the GNB/LNB pathway. These results suggest that GalT1 plays a more significant role than previously thought and predominates when B. bifidum grows on lactose and human milk oligosaccharides. GalT2 activity is required only during growth on substrates with a GNB core such as mucin glycans.

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References

Hidaka M, Nishimoto M, Kitaoka M, Wakagi T, Shoun H, Fushinobu S . The crystal structure of galacto-N-biose/lacto-N-biose I phosphorylase: a large deformation of a TIM barrel scaffold. J Biol Chem. 2009; 284(11):7273-83. PMC: 2652316. DOI: 10.1074/jbc.M808525200. View

Lee L, Kimura A, Tochikura T . Purification and properties of UDP-glucose (UDP-galactose) pyrophosphorylase from Bifidobacterium bifidum. J Biochem. 1979; 86(4):923-8. DOI: 10.1093/oxfordjournals.jbchem.a132624. View

Klaassens E, Ben-Amor K, Vriesema A, Vaughan E, de Vos W . The fecal bifidobacterial transcriptome of adults: a microarray approach. Gut Microbes. 2011; 2(4):217-26. DOI: 10.4161/gmic.2.4.16798. View

Mollet B, Pilloud N . Galactose utilization in Lactobacillus helveticus: isolation and characterization of the galactokinase (galK) and galactose-1-phosphate uridyl transferase (galT) genes. J Bacteriol. 1991; 173(14):4464-73. PMC: 208110. DOI: 10.1128/jb.173.14.4464-4473.1991. View

Geeganage S, Frey P . Significance of metal ions in galactose-1-phosphate uridylyltransferase: an essential structural zinc and a nonessential structural iron. Biochemistry. 1999; 38(40):13398-406. DOI: 10.1021/bi9910631. View

Sela D, Chapman J, Adeuya A, Kim J, Chen F, Whitehead T . The genome sequence of Bifidobacterium longum subsp. infantis reveals adaptations for milk utilization within the infant microbiome. Proc Natl Acad Sci U S A. 2008; 105(48):18964-9. PMC: 2596198. DOI: 10.1073/pnas.0809584105. View

Muthana M, Qu J, Li Y, Zhang L, Yu H, Ding L . Efficient one-pot multienzyme synthesis of UDP-sugars using a promiscuous UDP-sugar pyrophosphorylase from Bifidobacterium longum (BLUSP). Chem Commun (Camb). 2012; 48(21):2728-30. DOI: 10.1039/c2cc17577k. View

Gonzalez R, Klaassens E, Malinen E, de Vos W, Vaughan E . Differential transcriptional response of Bifidobacterium longum to human milk, formula milk, and galactooligosaccharide. Appl Environ Microbiol. 2008; 74(15):4686-94. PMC: 2519361. DOI: 10.1128/AEM.00122-08. View

Lee L, Kimura A, Tochikura T . Presence of a single enzyme catalyzing the pyrophosphorolysis of UDP-glucose and UDP-galactose in Bifidobacterium bifidum. Biochim Biophys Acta. 1978; 527(1):301-4. DOI: 10.1016/0005-2744(78)90281-4. View

10.

Datsenko K, Wanner B . One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci U S A. 2000; 97(12):6640-5. PMC: 18686. DOI: 10.1073/pnas.120163297. View

11.

Wu Z, Ethen C, Prather B, Machacek M, Jiang W . Universal phosphatase-coupled glycosyltransferase assay. Glycobiology. 2010; 21(6):727-33. DOI: 10.1093/glycob/cwq187. View

12.

Schell M, Karmirantzou M, Snel B, Vilanova D, Berger B, Pessi G . The genome sequence of Bifidobacterium longum reflects its adaptation to the human gastrointestinal tract. Proc Natl Acad Sci U S A. 2002; 99(22):14422-7. PMC: 137899. DOI: 10.1073/pnas.212527599. View

13.

Jensen L, Kuhn M, Stark M, Chaffron S, Creevey C, Muller J . STRING 8--a global view on proteins and their functional interactions in 630 organisms. Nucleic Acids Res. 2008; 37(Database issue):D412-6. PMC: 2686466. DOI: 10.1093/nar/gkn760. View

14.

Ruas-Madiedo P, Gueimonde M, Fernandez-Garcia M, de Los Reyes-Gavilan C, Margolles A . Mucin degradation by Bifidobacterium strains isolated from the human intestinal microbiota. Appl Environ Microbiol. 2008; 74(6):1936-40. PMC: 2268317. DOI: 10.1128/AEM.02509-07. View

15.

Kotake T, Yamaguchi D, Ohzono H, Hojo S, Kaneko S, Ishida H . UDP-sugar pyrophosphorylase with broad substrate specificity toward various monosaccharide 1-phosphates from pea sprouts. J Biol Chem. 2004; 279(44):45728-36. DOI: 10.1074/jbc.M408716200. View

16.

Frey P . The Leloir pathway: a mechanistic imperative for three enzymes to change the stereochemical configuration of a single carbon in galactose. FASEB J. 1996; 10(4):461-70. View

17.

Guarner F, Malagelada J . Gut flora in health and disease. Lancet. 2003; 361(9356):512-9. DOI: 10.1016/S0140-6736(03)12489-0. View

18.

Kitaoka M . Bifidobacterial enzymes involved in the metabolism of human milk oligosaccharides. Adv Nutr. 2012; 3(3):422S-9S. PMC: 3649479. DOI: 10.3945/an.111.001420. View

19.

Bradford M . A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976; 72:248-54. DOI: 10.1016/0003-2697(76)90527-3. View

20.

Asakuma S, Hatakeyama E, Urashima T, Yoshida E, Katayama T, Yamamoto K . Physiology of consumption of human milk oligosaccharides by infant gut-associated bifidobacteria. J Biol Chem. 2011; 286(40):34583-92. PMC: 3186357. DOI: 10.1074/jbc.M111.248138. View