The effect of N-terminal truncation on double-dimer assembly of goose delta-crystallin.
Delta-crystallin is a soluble structural protein in avian eye lenses that confers special refractive properties. In the presence of GdmCl (guanidinium chloride), tetrameric delta-crystallin undergoes dissociation via a dimeric state to a monomeric molten globule intermediate state. The latter are denatured at higher GdmCl concentrations in a multi-state manner. In the present study, the X-ray structure of goose delta-crystallin was determined to 2.8 A (1 A=0.1 nm). In this structure the first 25 N-terminal residues interact with a hydrophobic cavity in a neighbouring molecule, stabilizing the quaternary structure of this protein. When these 25 residues were deleted this did not produce any gross structural changes, as judged by CD analysis, but slightly altered tryptophan fluorescence and ANS (8-anilino-1-naphthalenesulphonic acid) spectra. The dimeric form was significantly identified as judged by sedimentation velocity and nondenaturing gradient gel electrophoresis. This mutant had increased sensitivity to temperature denaturation and GdmCl concentrations of 0.3-1.0 M. This protein was destabilized about 3.3 kcal/mol (1 kcal=4.184 kJ) due to N-terminal truncation. After incubation at 37 degrees C N-terminal truncated proteins were prone to aggregation, suggesting the presence of the unstable dimeric conformation. An important role for the N-terminus in dimer assembly of goose delta-crystallin is proposed.
Lee
Hwei-Jen
HJ
Department of Biochemistry, National Defense Medical Center, Taipei 114, Taiwan. hjlee@mail.ndmctsgh.edu.tw
Lai
Young-Hsang
YH
Wu
Su-Ying
SY
Chen
Yu-Hou
YH
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D005777:Geese; D019791:Guanidine / Q000737:chemistry / Q000494:pharmacology; D008958:Models, Molecular; D017510:Protein Folding; D020836:Protein Structure, Quaternary / Q000187:drug effects; D017384:Sequence Deletion / Q000235:genetics*; D013379:Substrate Specificity; D013696:Temperature; D038207:delta-Crystallins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism
D038207:delta-Crystallins; D019791:Guanidine