Abstract
We have used total chemical synthesis to prepare [V15A]crambin-α- carboxamide, a unique protein analogue that eliminates a salt bridge between the δ-guanidinium of the Arg10 side chain and the α-carboxylate of Asn46 at the C-terminus of the polypeptide chain. This salt bridge is thought to be important for the folding and stability of the crambin protein molecule. Folding, with concomitant disulfide bond formation, of the fully reduced [V15A]crambin-α-carboxamide polypeptide was less efficient than folding/disulfide formation for the [V15A]crambin polypeptide under a standard set of conditions. To probe the origin of this less efficient folding/disulfide bond formation, we separately crystallized purified synthetic [V15A]crambin-α-carboxamide and chemically synthesized [V15A]crambin and solved their X-ray structures. The crystal structure of [V15A]crambin-α-carboxamide showed that elimination of the Arg 10-Asn46 salt bridge caused disorder of the C-terminal region of the polypeptide chain and affected the overall 'tightness' of the structure of the protein molecule. These studies, enabled by chemical protein synthesis, strongly suggest that in native crambin the Arg10-Asn 46 salt bridge contributes to efficient formation of correct disulfide bonds and also to the well-ordered structure of the protein molecule.
| Original language | English |
|---|---|
| Pages (from-to) | 750-756 |
| Number of pages | 7 |
| Journal | Molecular BioSystems |
| Volume | 5 |
| Issue number | 7 |
| DOIs | |
| Publication status | Published - 2009 Jul 20 |
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All Science Journal Classification (ASJC) codes
- Biotechnology
- Molecular Biology
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Role of a salt bridge in the model protein crambin explored by chemical protein synthesis : X-ray structure of a unique protein analogue, [V15A]crambin-α-carboxamide. / Bang, Duhee; Tereshko, Valentina; Kossiakoff, Anthony A.; Kent, Stephen B.H.
In: Molecular BioSystems, Vol. 5, No. 7, 20.07.2009, p. 750-756.Research output: Contribution to journal › Article
TY - JOUR
T1 - Role of a salt bridge in the model protein crambin explored by chemical protein synthesis
T2 - X-ray structure of a unique protein analogue, [V15A]crambin-α-carboxamide
AU - Bang, Duhee
AU - Tereshko, Valentina
AU - Kossiakoff, Anthony A.
AU - Kent, Stephen B.H.
PY - 2009/7/20
Y1 - 2009/7/20
N2 - We have used total chemical synthesis to prepare [V15A]crambin-α- carboxamide, a unique protein analogue that eliminates a salt bridge between the δ-guanidinium of the Arg10 side chain and the α-carboxylate of Asn46 at the C-terminus of the polypeptide chain. This salt bridge is thought to be important for the folding and stability of the crambin protein molecule. Folding, with concomitant disulfide bond formation, of the fully reduced [V15A]crambin-α-carboxamide polypeptide was less efficient than folding/disulfide formation for the [V15A]crambin polypeptide under a standard set of conditions. To probe the origin of this less efficient folding/disulfide bond formation, we separately crystallized purified synthetic [V15A]crambin-α-carboxamide and chemically synthesized [V15A]crambin and solved their X-ray structures. The crystal structure of [V15A]crambin-α-carboxamide showed that elimination of the Arg 10-Asn46 salt bridge caused disorder of the C-terminal region of the polypeptide chain and affected the overall 'tightness' of the structure of the protein molecule. These studies, enabled by chemical protein synthesis, strongly suggest that in native crambin the Arg10-Asn 46 salt bridge contributes to efficient formation of correct disulfide bonds and also to the well-ordered structure of the protein molecule.
AB - We have used total chemical synthesis to prepare [V15A]crambin-α- carboxamide, a unique protein analogue that eliminates a salt bridge between the δ-guanidinium of the Arg10 side chain and the α-carboxylate of Asn46 at the C-terminus of the polypeptide chain. This salt bridge is thought to be important for the folding and stability of the crambin protein molecule. Folding, with concomitant disulfide bond formation, of the fully reduced [V15A]crambin-α-carboxamide polypeptide was less efficient than folding/disulfide formation for the [V15A]crambin polypeptide under a standard set of conditions. To probe the origin of this less efficient folding/disulfide bond formation, we separately crystallized purified synthetic [V15A]crambin-α-carboxamide and chemically synthesized [V15A]crambin and solved their X-ray structures. The crystal structure of [V15A]crambin-α-carboxamide showed that elimination of the Arg 10-Asn46 salt bridge caused disorder of the C-terminal region of the polypeptide chain and affected the overall 'tightness' of the structure of the protein molecule. These studies, enabled by chemical protein synthesis, strongly suggest that in native crambin the Arg10-Asn 46 salt bridge contributes to efficient formation of correct disulfide bonds and also to the well-ordered structure of the protein molecule.
UR - http://www.scopus.com/inward/record.url?scp=67650367214&partnerID=8YFLogxK
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U2 - 10.1039/b903610e
DO - 10.1039/b903610e
M3 - Article
C2 - 19562114
AN - SCOPUS:67650367214
VL - 5
SP - 750
EP - 756
JO - Molecular BioSystems
JF - Molecular BioSystems
SN - 1742-206X
IS - 7
ER -