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Biochemical Characterization of the Pneumococcal Glucose 1-Phosphate Uridylyltransferase (GalU) Essential for Capsule Biosynthesis

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Abstract

The glucose 1-phosphate uridylyltransferase (GalU) is absolutely required for the biosynthesis of capsular polysaccharide, the sine qua non virulence factor of Streptococcus pneumoniae. The pneumococcal GalU protein was overexpressed in Escherichia coli, and purified. GalU showed a pI of 4.23, and catalyzed the reversible formation of UDP-glucose and pyrophosphate from UTP and glucose 1-phosphate with Km values of 0.4 mM for UDP-glucose, 0.26 mM for pyrophosphate, 0.19 mM for glucose 1-phosphate, and 0.24 mM for UTP. GalU has an optimum pH of 8–8.5, and requires Mg2+ for activity. Neither ADP-glucose nor TDP-glucose is utilized as substrates in vitro. The purification of GalU represents a fundamental step to provide insights on drug design to control the biosynthesis of the main pneumococcal virulence factor.

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Acknowledgments

We thank R. López and P. García for a critical reading of the manuscript. This work was supported by grants from Universidad de Buenos Aires, Agencia National de Promoción Científica y Tecnológica (Argentina), Dirección General de Investigatión Científica y Técnica (BCM2003-00074), and Redes Temáticas de Investigación Cooperativa (G03/103 and C03/14) (Ministerio de Sanidad y Consumo Spain).

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Authors and Affiliations

  1. Cátedra de Microbiología, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, , Junín 956, , 1113, Buenos Aires, Argentina

    Laura Bonofiglio & Marta Mollerach

  2. Centro de Investigaciones Biólogicas, CSIC, Ramiro de Maetzu 9, 28040 , Madrid, Spain

    Ernesto García

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  1. Laura Bonofiglio
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  2. Ernesto García
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  3. Marta Mollerach
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Correspondence to Marta Mollerach.

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Bonofiglio, L., García, E. & Mollerach, M. Biochemical Characterization of the Pneumococcal Glucose 1-Phosphate Uridylyltransferase (GalU) Essential for Capsule Biosynthesis. Curr Microbiol 51, 217–221 (2005). https://doi.org/10.1007/s00284-005-4466-0

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