A glycosyltransferase with a length-controlling activity as a mechanism to regulate the size of polysaccharides

doi:10.1073/pnas.0708025104

A glycosyltransferase with a length-controlling activity as a mechanism to regulate the size of polysaccharides

*Instituto de Investigaciones Biotecnológicas, Universidad Nacional de General San Martín, 1650 Buenos Aires, Argentina; and
^†Centro de Investigaciones en Hidratos de Carbono–Consejo Nacional de Investigaciones Científicas y Técnicas de Argentina, Departamento de Química Orgánica, Facultad de Ciencias Exactas y Naturales, Universidad de Buenos Aires, 2160 Ciudad Universitaria, Argentina

Communicated by Armando J. Parodi, Fundación Instituto Leloir, Buenos Aires, Argentina, August 30, 2007 (received for review May 28, 2007)

Abstract

Cyclic β-1,2-glucans (CβG) are osmolyte homopolysaccharides with a cyclic β-1,2-backbone of 17–25 glucose residues present in the periplasmic space of several bacteria. Initiation, elongation, and cyclization, the three distinctive reactions required for building the cyclic structure, are catalyzed by the same protein, the CβG synthase. The initiation activity catalyzes the transference of the first glucose from UDP-glucose to a yet-unidentified amino acid residue in the same protein. Elongation proceeds by the successive addition of glucose residues from UDP-glucose to the nonreducing end of the protein-linked β-1,2-oligosaccharide intermediate. Finally, the protein-linked intermediate is cyclized, and the cyclic glucan is released from the protein. These reactions do not explain, however, the mechanism by which the number of glucose residues in the cyclic structure is controlled. We now report that control of the degree of polymerization (DP) is carried out by a β-1,2-glucan phosphorylase present at the CβG synthase C-terminal domain. This last activity catalyzes the phosphorolysis of the β-1,2-glucosidic bond at the nonreducing end of the linear protein-linked intermediate, releasing glucose 1-phosphate. The DP is thus regulated by this “length-controlling” phosphorylase activity. To our knowledge, this is the first description of a control of the DP of homopolysaccharides.

Footnotes

^‡To whom correspondence should be addressed at:
Instituto de Investigaciones Biotecnológicas, Universidad Nacional de General San Martín, Instituto Nacional de Tecnologia Industrial, Edifício 24, Avenida General Paz 5445, San Martín, 1650 Buenos Aires, Argentina.
E-mail: rugalde{at}iib.unsam.edu.ar
Author contributions: A.E.C., L.S.G., N.I.d.I., and R.A.U. designed research; A.E.C. and L.S.G. performed research; A.C.C. and A.S.C. contributed new reagents/analytic tools; A.E.C., L.S.G., A.C.C., A.S.C., N.I.d.I., and R.A.U. analyzed data; and A.E.C. and R.A.U. wrote the paper.
The authors declare no conflict of interest.
This article contains supporting information online at www.pnas.org/cgi/content/full/0708025104/DC1.
Abbreviations:

CBM,

carbohydrate-binding module;

ChBP,

chitobiose phosphorylase;

CβG,

cyclic β-1,2-glucans;

Cgs,

CβG synthase;

DP,

degree of polymerization;

glucose-1-P,

glucose 1-phosphate;

GT,

glycosyltransferase;

HPAEC,

high-pH anion-exchange chromatography;

PAD,

pulse amperometric detection;

TMS,

transmembrane-spanning segment.