title: Structural study of the G57W mutant of human gamma-S-crystallin, associated with congenital cataract.
abstract: Human γS-crystallin (CrygS) is an important component of the human eye lens nucleus and cortex. The mutation G57W in the molecule is reported to be associated with congenital cataract in children. We compare the conformational features and aggregation properties of the mutant protein G57W with the wild-type CrygS to understand how the structural changes in the mutant are related to the mechanism of opacification.
  
  
  
  Wild-type and mutant proteins were cloned, expressed, and purified, and their structural properties were studied in solution. Conformational features and the structural stability of the proteins were compared in solution, using circular dichroism (CD) and fluorescence spectroscopic analysis, and the proteins' tendencies to aggregate were compared using extrinsic spectral probes. In addition, we analyzed the proteins' structural differences with extensive molecular modeling in silico.
  
  
  
  CD and intrinsic fluorescence analysis suggested the secondary and tertiary structures of the mutant are slightly altered. Experiments using extrinsic spectral probes revealed that the compact close-packed structure is loosened somewhat, and the mutant tends to self-aggregate. Denaturation (both thermal and chemical) studies indicate that the replacement of glycine (G) in position 57 by tryptophan (W) lowered the structural stability of the molecule. Further, the mutant had a tendency to precipitate and scatters light more easily than the wild-type.
  
  
  
  The replacement of glycine at position 57 by the tryptophan residue in human γS-crystallin weakens the stability of the mutant molecule and causes the molecule to self-aggregate, thus generating light-scattering particles. This set of changes in the mutant offers a molecular insight into the mechanism of opacification.
authors.affiliation: Prof. Brien Holden Eye Research Centre, Hyderabad Eye Research Foundation, L. V. Prasad Eye Institute, Hyderabad, Telangana, India.
authors.forename: Ismail
authors.initials: I
authors.lastname: Khan
authors.affiliation: Plot 32, LIC Colony,W Marredpally, Secunderabad, Telangana, India.
authors.forename: Sushil
authors.initials: S
authors.lastname: Chandani
authors.affiliation: Prof. Brien Holden Eye Research Centre, Hyderabad Eye Research Foundation, L. V. Prasad Eye Institute, Hyderabad, Telangana, India.
authors.forename: Dorairajan
authors.initials: D
authors.lastname: Balasubramanian
mesh_terms: D019943:Amino Acid Substitution; D002386:Cataract / Q000151:congenital*; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D006801:Humans; D008958:Models, Molecular; D050505:Mutant Proteins / Q000737:chemistry*; D017354:Point Mutation; D066329:Protein Aggregates; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D013050:Spectrometry, Fluorescence; D038222:gamma-Crystallins / Q000737:chemistry* / Q000235:genetics*
chemical_list: D050505:Mutant Proteins; D066329:Protein Aggregates; D038222:gamma-Crystallins; C077648:CRYGS protein, human