IBR   13079
INSTITUTO DE BIOLOGIA MOLECULAR Y CELULAR DE ROSARIO
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Distinctive properties of ADP-glucose pyrophosphorylases from Gram-positive bacteria
Autor/es:
ASENCIÓN DIEZ, MATÍAS D.; DEMONTE, ANA MARÍA; PEIRÚ, SALVADOR; GRAMAJO, HUGO; GUERRERO, SERGIO A.; IGLESIAS, ALBERTO A.
Lugar:
San Miguel de Tucumán
Reunión:
Congreso; VII Congreso de Microbiología General SAMIGE del Bicentenario; 2011
Institución organizadora:
Sociedad Argentina de Microbiología General -SAMIGE-
Resumen:
Glycogen or similar
α-1,4-polyglucans were reported in more than 50 different bacterial
species. This is not restricted to any class of bacteria, as many Gram-negative
and Gram-positive bacteria as well as archaebacteria have been reported to accumulate
the polysaccharide. The process occurs
through ADP‑glucose (ADPGlc) as glucosyl donor for elongation of the
polyglucan. ADPGlc is produced via
ADPGlc pyrophosphorylase (ADPGlcPPase, EC 2.7.7.27) in a key regulatory step of
the biosynthetic pathway. ADPGlcPPase has been purified from a number of
microorganisms, and as a rule is was found homotetrameric in structure. The one
exception is the enzyme from <i>Bacillus</i> sp., as in <i>B. stearothermophilus</i> it is also
a tetramer, but composed of two subunits: GlgC (active) and GlgD. Also,
ADPGlcPPases from bacteria are allosterically regulated enzymes. In general,
ADPGlcPPase effectors are main metabolites of the major pathway for carbon
assimilation in the respective organism.
Studies of enzymes involved
in glycogen metabolism in Gram‑positive bacteria are scarce, which represents
an important deficit for the understanding of carbon metabolism in these
organisms. We characterized the kinetic, regulatory and structural properties
of ADPGlcPPases from <i>Mycobacterium tuberculosis</i>
(<i>Mtu</i>), <i>Streptomyces coelicolor</i>
(<i>Sco</i>) and <i>Streptococcus mutans</i>
(<i>Smu</i>). <i>Mtu</i> and <i>Sco</i>
ADPGlcPPases share 60% identity at the protein level and both were activated by
PEP and Glc6P, the latter being a novel activator for bacterial ADPGlcPPases.
In contrast, we found differences concerning affinity for substrates and enzyme
activity, as well as sensitivity to allosteric effectors. Particularly, the
<i>Sco</i> enzyme is 20-fold less active than the
<i>Mtu</i> counterpart, but the former has 5-fold more affinity for
substrates and exhibits more promiscuity to effectors, being also inhibited by
NADPH and Pi.
Concerning the
<i>Smu</i> ADPGlcPPase, it is composed by two subunits: GlgC and
GlgD [this being a characteristic of the enzyme from firmicutes], and we
determined that GlgC, but not GlgD, is the catalytic one. Also, the heteromeric
conformation GlgCD has activity one
order of magnitude higher than homomeric GlgC and both conformations
have distinctive allosteric effectors.
Fru-1,6-bisP activated 2-fold GlgC activity, but it was ineffective on GlgCD.
As well, PEP inhibited GlgCD but not GlgC activity. Our results represent key
differences regarding the enzyme from <i>B. stearothermophilus</i>,
which is an unregulated ADPGlcPPase exhibiting no difference in
<i>V</i><sub>max</sub> between GlgC and GlgCD
conformations.
The whole view
supports that regulation of the key regulatory enzyme
for glycogen synthesis in Gram-positive bacteria shows peculiarities.
These seem closely related to particular metabolic routes in the respective
organism operating in carbon and energy accumulation as well as in Glc1P
partitioning.