Expression of the partial protein encoded by mutated COL4A5 gene and analysis of the structure by circular dichroism
- VernacularTitle:突变型COL4A5基因编码蛋白片段的表达及圆二色谱结构分析
- Author:
Yunfeng WANG
;
Jie DING
;
Dingfang BU
;
Fang WANG
;
- Publication Type:Journal Article
- Keywords:
Nephritis, hereditary/genet;
Mutation;
Collagen;
Structure
- From:
Journal of Peking University(Health Sciences)
2004;0(02):-
- CountryChina
- Language:Chinese
-
Abstract:
T mutation resulting in p.G1015V from an X linked Alport patient, and that of corresponding cDNA from a control were expressed in E.coli. The recombinant and mutant polypeptide was a fragment of COL4A5 , containing 158 amino acid residues with a glycine to valine substitution mutation in it. The secondary structure of the two recombinant proteins was analyzed using circular dichroism(CD) spectroscopy. Results: CD spectra of the control exhibited a negative peak near 200 nm whereas that of the patient exhibited a negative peak near 220 nm. The magnitude of the negative peak of the patient decreased as compared with that of the control. Furthermore, secondary structure of the control polypeptide was mainly composed of ? sheet and random coil without ? helix, whereas that of the patient presented 12.9% ? helix. Conclusion: Not only local structure of the substitution site but also folding kinetics of the entire ?5 chain may be changed due to Gly→Val substitution in Alport syndrome. We speculate that the abnormally folded polypeptide chain may not be assembled into the triple helix and the network of type Ⅳ collagen, or may be assembled into loosen triple helix then degraded easily, resulting in the pathognomonic ultrastructural changes of the glomerular basement membrane.
