 |
PDBsum entry 3brm
|
|
|
|
References listed in PDB file
|
 |
|
Key reference
|
|
|
Title
|
|
Functional and structural characterization of four glutaminases from escherichia coli and bacillus subtilis.
|
|
|
Authors
|
|
G.Brown,
A.Singer,
M.Proudfoot,
T.Skarina,
Y.Kim,
C.Chang,
I.Dementieva,
E.Kuznetsova,
C.F.Gonzalez,
A.Joachimiak,
A.Savchenko,
A.F.Yakunin.
|
|
|
Ref.
|
|
Biochemistry, 2008,
47,
5724-5735.
|
|
|
PubMed id
|
|
|
|
|
|
Abstract
|
|
|
Glutaminases belong to the large superfamily of serine-dependent beta-lactamases
and penicillin-binding proteins, and they catalyze the hydrolytic deamidation of
L-glutamine to L-glutamate. In this work, we purified and biochemically
characterized four predicted glutaminases from Escherichia coli (YbaS and YneH)
and Bacillus subtilis (YlaM and YbgJ). The proteins demonstrated strict
specificity to L-glutamine and did not hydrolyze D-glutamine or L-asparagine. In
each organism, one glutaminase showed higher affinity to glutamine ( E. coli
YbaS and B. subtilis YlaM; K m 7.3 and 7.6 mM, respectively) than the second
glutaminase ( E. coli YneH and B. subtilis YbgJ; K m 27.6 and 30.6 mM,
respectively). The crystal structures of the E. coli YbaS and the B. subtilis
YbgJ revealed the presence of a classical beta-lactamase-like fold and
conservation of several key catalytic residues of beta-lactamases (Ser74, Lys77,
Asn126, Lys268, and Ser269 in YbgJ). Alanine replacement mutagenesis
demonstrated that most of the conserved residues located in the putative
glutaminase catalytic site are essential for activity. The crystal structure of
the YbgJ complex with the glutaminase inhibitor 6-diazo-5-oxo- l-norleucine
revealed the presence of a covalent bond between the inhibitor and the hydroxyl
oxygen of Ser74, providing evidence that Ser74 is the primary catalytic
nucleophile and that the glutaminase reaction proceeds through formation of an
enzyme-glutamyl intermediate. Growth experiments with the E. coli glutaminase
deletion strains revealed that YneH is involved in the assimilation of
l-glutamine as a sole source of carbon and nitrogen and suggested that both
glutaminases (YbaS and YneH) also contribute to acid resistance in E. coli.
|
|
|
|
|
|
|
