TI: Sequencing of the ddl gene and modeling of the
mutated D-alanine:D-alanine ligase in glycopeptide-dependent strains of
Enterococcus faecium.
AU: Gholizadeh,-Y; Prevost,-M; Van-Bambeke,-F; Casadewall,-B;
Tulkens,-P-M; Courvalin,-P
AD: Unite des Agents Antibacteriens, Institut Pasteur, Paris,
France.
SO: Protein-Sci. 2001 Apr; 10(4): 836-44
JN: Protein-science
PY: 2001
AB: Glycopeptide dependence for growth in enterococci results
from mutations in the ddl gene that inactivate the host D-Ala:D-Ala ligase.
The strains require glycopeptides as inducers for synthesis of resistance
proteins, which allows for the production of peptidoglycan precursors ending
in D-Ala-D-Lac instead of D-Ala-D-Ala. The sequences of the ddl gene from
nine glycopeptide-dependent Enterococcus faecium clinical isolates were
determined. Each one had a mutation consisting either in a 5-bp insertion
at position 41 leading to an early stop codon, an in-frame 6-bp deletion
causing the loss of two residues (KDVA243-246 to KA), or single base-pair
changes resulting in an amino acid substitution (E13 --> G, G99 --> R,
V241 --> D, D295 --> G, P313 --> L). The potential consequences of the
deletion and point mutations on the 3-D structure of the enzyme were evaluated
by comparative molecular modeling of the E. faecium enzyme, using the X-ray
structure of the homologous Escherichia coli D-Ala:D-Ala ligase DdlB as
a template. All mutated residues were found either to interact directly
with one of the substrates of the enzymatic reaction (E13 and D295) or
to stabilize the position of critical residues in the active site. Maintenance
of the 3-D structure in the vicinity of these mutations in the active site
appears critical for D-Ala:D-Ala ligase activity.