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PDBsum entry 4ha2
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Hydrolase inhibitor
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PDB id
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4ha2
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References listed in PDB file
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Key reference
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Title
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A conserved tryptophan (w91) at the barrel-Lid junction modulates the packing and stability of kunitz (sti) family of inhibitors.
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Authors
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S.Majumder,
S.Khamrui,
R.Banerjee,
P.Bhowmik,
U.Sen.
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Ref.
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Biochim Biophys Acta, 2015,
1854,
55-64.
[DOI no: ]
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PubMed id
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Abstract
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β-trefoil fold, consisting of a six stranded β-barrel capped at one end by a
lid comprising of another six β-strands, is one of the most important folds
among proteins. Important classes of proteins like Interleukins (ILs),
Fibroblast Growth Factors (FGFs), Kunitz (STI) family of inhibitors etc. belong
to this fold. Their core is packed by hydrophobic residues contributed by the 6
stranded β-barrel and three β-hairpins that make essential contacts with each
other and keep the protein in 'topologically minimal frustrated state'. A
complete database analysis of the core residues of the β-trefoil fold proteins
presented here identified a conserved tryptophan (W91) residue in the Kunitz
(STI) family of inhibitors that projects from the lid and interacts with the
bottom layer residues of the barrel. This kind of interactions is unique in
Kunitz (STI) family because no other families of β-trefoil fold have such a
shear sized residue at the barrel lid junction; suggesting its possible
importance in packing and stability. We took WCI as a representative of this
family and prepared four cavity creating mutants W91F-WCI, W91M-WCI, W91I-WCI
& W91A-WCI. CD experiments show that the secondary structure of the mutants
remains indistinguishable with the wild type. Crystal structures of the mutants
W91F-WCI, W91M-WCI & W91A-WCI also show the same feature. However, slight
readjustments of the side chains around the site of mutation have been observed
so as to minimize the cavity created due to mutation. Comparative stability of
these mutants, estimated using heat denaturation CD spectroscopy, indicates that
stability of the mutants inversely correlates with the size of the cavity inside
the core. Interestingly, although we mutated at the core, mutants show varying
susceptibility against tryptic digestion that grossly follow their instability
determined by CD. Our findings suggest that the W91 residue plays an important
role in determining the stability and packing of the core of WCI.
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