 |
PDBsum entry 2c0r
|
|
|
|
References listed in PDB file
|
 |
|
Key reference
|
|
|
Title
|
|
Effect of ph on the structure and stability of bacillus circulans ssp. Alkalophilus phosphoserine aminotransferase: thermodynamic and crystallographic studies.
|
|
|
Authors
|
|
E.G.Kapetaniou,
A.Thanassoulas,
A.P.Dubnovitsky,
G.Nounesis,
A.C.Papageorgiou.
|
|
|
Ref.
|
|
Proteins, 2006,
63,
742-753.
[DOI no: ]
|
|
|
PubMed id
|
|
|
|
|
Note In the PDB file this reference is
annotated as "TO BE PUBLISHED".
The citation details given above were identified by an automated
search of PubMed on title and author
names, giving a
perfect match.
|
|
|
|
Abstract
|
|
|
pH is one of the key parameters that affect the stability and function of
proteins. We have studied the effect of pH on the
pyridoxal-5'-phosphate-dependent enzyme phosphoserine aminotransferase produced
by the facultative alkaliphile Bacillus circulans ssp. alkalophilus using
thermodynamic and crystallographic analysis. Enzymatic activity assay showed
that the enzyme has maximum activity at pH 9.0 and relative activity less than
10% at pH 7.0. Differential scanning calorimetry and circular dichroism
experiments revealed variations in the stability and denaturation profiles of
the enzyme at different pHs. Most importantly, release of pyridoxal-5'-phosphate
and protein thermal denaturation were found to occur simultaneously at pH 6.0 in
contrast to pH 8.5 where denaturation preceded cofactor's release by
approximately 3 degrees C. To correlate the observed differences in thermal
denaturation with structural features, the crystal structure of phosphoserine
aminotransferase was determined at 1.2 and 1.5 A resolution at two different pHs
(8.5 and 4.6, respectively). Analysis of the two structures revealed changes in
the vicinity of the active site and in surface residues. A conformational change
in a loop involved in substrate binding at the entrance of the active site has
been identified upon pH change. Moreover, the number of intramolecular ion pairs
was found reduced in the pH 4.6 structure. Taken together, the presented
kinetics, thermal denaturation, and crystallographic data demonstrate a
potential role of the active site in unfolding and suggest that subtle but
structurally significant conformational rearrangements are involved in the
stability and integrity of phosphoserine aminotransferase in response to pH
changes.
|
|
|
|
|
|
|
|
Figure 1.
Figure 1. Schematic representation of the chemical reaction
catalyzed by phosphoserine aminotransferase.
|
|
Figure 7.
Figure 7. Ribbon representation of the BCIR PSAT structure at
pH 8.5 (A) dimer (B) monomer. Secondary-structure elements were
calculated using DSSP.[29] The  -helices
are shown in yellow and the  -strands
in cyan. N- and C-termini are shown as spheres and PLP molecules
in space filling (A) and ball-and-stick (B). The small domain
contains an extended five-stranded -sheet,
composed of a two-stranded parallel -sheet
( 1
and 13)
and a three-stranded antiparallel -sheet
(+ 11,
- 12,
- 14).
The large domain contains the conservative among PLP-dependent
enzymes[44] seven-stranded -sheet
( 2-
5
and 7-
9)
with all -strands
parallel except 9.
Two additional -strands
( 6
and 10),
six -helices
( 1-
6),
and six 3[10]-helices complete the large domain. Helix 7
connects the large domain with the small domain. The
five-stranded -sheet
is flanked by helix 7
and two additional helices ( 8
and 9)
close to the C-terminus.
|
|
|
|
|
|
The above figures are
reprinted
by permission from John Wiley & Sons, Inc.:
Proteins
(2006,
63,
742-753)
copyright 2006.
|
|
|
|
|
|
|
