Characterization of denatured proteins by hydrophobic interaction chromatography: a preliminary study.
In this preliminary study hydrophobic interaction chromatography (HIC) is proposed as a good tool in order to detect conformational changes induced by chemical denaturants in two globular proteins, cytochrome C (Cyt C) and myoglobin (MYO). Alterations in protein structure were manifested chromatographically by reproducible changes in peak heights, retention time, and appearance of multiple peaks. The HIC behavior of the two model proteins denatured by guanidinium thyocyanate (GdmSCN) was investigated, keeping constant various concentrations of urea in the mobile phase in a TSK-Gel Phenyl-5PW column (TosoBiosep). Suitable elution conditions provide evidence of the simultaneous presence of two denatured forms in the case of MYO, and sequential different denatured states of Cyt C.
Bramanti
Emilia
E
National Council of Research-CNR, Istituto per i Processi Chimico-Fisici, Laboratory of Instrumental Analytical Chemistry, Via G. Moruzzi 1, 56124-Pisa, Italy. emilia@ipcf.cnr.it
Ferri
Fabrizio
F
Sortino
Chandra
C
Onor
Massimo
M
Raspi
Giorgio
G
Venturini
Marcella
M
D000818:Animals; D002021:Buffers; D002845:Chromatography / Q000379:methods*; D045304:Cytochromes c / Q000737:chemistry; D006146:Guanidines / Q000494:pharmacology; D006736:Horses; D057927:Hydrophobic and Hydrophilic Interactions; D008958:Models, Molecular; D009211:Myoglobin / Q000737:chemistry; D011489:Protein Denaturation / Q000187:drug effects; D011506:Proteins / Q000737:chemistry*; D012492:Salts / Q000737:chemistry; D013861:Thiocyanates / Q000494:pharmacology; D014508:Urea / Q000737:chemistry / Q000494:pharmacology
D002021:Buffers; D006146:Guanidines; D009211:Myoglobin; D011506:Proteins; D012492:Salts; D013861:Thiocyanates; C054436:guanidine thiocyanate; D014508:Urea; D045304:Cytochromes c