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PDBsum entry 2htx
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Contents |
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* Residue conservation analysis
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Enzyme class:
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E.C.3.2.1.17
- lysozyme.
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Reaction:
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Hydrolysis of the 1,4-beta-linkages between N-acetyl-D-glucosamine and N-acetylmuramic acid in peptidoglycan heteropolymers of the prokaryotes cell walls.
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Biotechnol Bioeng
98:711-718
(2007)
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PubMed id:
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Elucidation of the mechanism and end products of glutaraldehyde crosslinking reaction by X-ray structure analysis.
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Y.Wine,
N.Cohen-Hadar,
A.Freeman,
F.Frolow.
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ABSTRACT
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Glutaraldehyde has been used for several decades as an effective crosslinking
agent for many applications including sample fixation for microscopy, enzyme and
cell immobilization, and stabilization of protein crystals. Despite of its
common use as a crosslinking agent, the mechanism and chemistry involved in
glutaraldehyde crosslinking reaction is not yet fully understood. Here we
describe feasibility study and results obtained from a new approach to
investigate the process of protein crystals stabilization by glutaraldehyde
crosslinking. It involves exposure of a model protein crystal (Lysozyme) to
glutaraldehyde in alkaline or acidic pH for different incubation periods and
reaction arrest by medium exchange with crystallization medium to remove unbound
glutaraldehyde. The crystals were subsequently incubated in diluted buffer
affecting dissolution of un-crosslinked crystals. Samples from the resulting
solution were subjected to protein composition analysis by gel electrophoresis
and mass spectroscopy while crosslinked, dissolution resistant crystals were
subjected to high resolution X-ray structural analysis. Data from gel
electrophoresis indicated that the crosslinking process starts at specific
preferable crosslinking site by lysozyme dimer formation, for both acidic and
alkaline pH values. These dimer formations were followed by trimer and tetramer
formations leading eventually to dissolution resistant crystals. The
crosslinking initiation site and the end products obtained from glutaraldehyde
crosslinking in both pH ranges resulted from reactions between lysine residues
of neighboring protein molecules and the polymeric form of glutaraldehyde.
Reaction rate was much faster at alkaline pH. Different reaction end products,
indicating different reaction mechanisms, were identified for crosslinking
taking place under alkaline or acidic conditions.
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Literature references that cite this PDB file's key reference
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PubMed id
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Reference
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N.Cohen-Hadar,
S.Lagziel-Simis,
Y.Wine,
F.Frolow,
and
A.Freeman
(2011).
Re-structuring protein crystals porosity for biotemplating by chemical modification of lysine residues.
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Biotechnol Bioeng,
108,
1.
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M.Salin,
E.G.Kapetaniou,
M.Vaismaa,
M.Lajunen,
M.G.Casteleijn,
P.Neubauer,
L.Salmon,
and
R.K.Wierenga
(2010).
Crystallographic binding studies with an engineered monomeric variant of triosephosphate isomerase.
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Acta Crystallogr D Biol Crystallogr,
66,
934-944.
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PDB codes:
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H.Koizumi,
M.Tachibana,
and
K.Kojima
(2009).
Elastic constants in tetragonal hen egg-white lysozyme crystals containing large amount of water.
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Phys Rev E Stat Nonlin Soft Matter Phys,
79,
061917.
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O.A.Andersen,
D.L.Schönfeld,
I.Toogood-Johnson,
B.Felicetti,
C.Albrecht,
T.Fryatt,
M.Whittaker,
D.Hallett,
and
J.Barker
(2009).
Cross-linking of protein crystals as an aid in the generation of binary protein-ligand crystal complexes, exemplified by the human PDE10a-papaverine structure.
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Acta Crystallogr D Biol Crystallogr,
65,
872-874.
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PDB code:
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W.Zhai,
J.Chang,
X.Lü,
and
Z.Wang
(2009).
Procyanidins-crosslinked heart valve matrix: anticalcification effect.
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J Biomed Mater Res B Appl Biomater,
90,
913-921.
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Y.Wine,
N.Cohen-Hadar,
R.Lamed,
A.Freeman,
and
F.Frolow
(2009).
Modification of protein crystal packing by systematic mutations of surface residues: implications on biotemplating and crystal porosity.
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Biotechnol Bioeng,
104,
444-457.
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A.Russo,
N.Chandramouli,
L.Zhang,
and
H.Deng
(2008).
Reductive glutaraldehydation of amine groups for identification of protein N-termini.
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J Proteome Res,
7,
4178-4182.
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The most recent references are shown first.
Citation data come partly from CiteXplore and partly
from an automated harvesting procedure. Note that this is likely to be
only a partial list as not all journals are covered by
either method. However, we are continually building up the citation data
so more and more references will be included with time.
Where a reference describes a PDB structure, the PDB
codes are
shown on the right.
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Links
