UV Raman monitoring of histidine protonation and H-(2)H exchange in plastocyanin.
J Inorg Biochem
; 88(3-4): 381-7, 2002 Feb.
Article
em En
| MEDLINE
| ID: mdl-11897354
ABSTRACT
UV resonance Raman bands of Cu-bound and protonated histidine residues have been detected in (2)H(2)O solutions of poplar plastocyanin. For the Cu(II) protein, slow NH-(2)H exchange of the His37 ligand was monitored via the growth of bands at 1389 and 1344 cm(-1) when Pcy was exchanged into (2)H(2)O, or via their diminution when the protein was exchanged back into H(2)O; the rate constant is 7 x 10(-4)/s at pH (p(2)H) 7.4 at room temperature. The slow exchange is attributed to imidazole H-bonding to a backbone carbonyl. Nearby bands at 1397 and 1354 cm(-1), appear and disappear within the mixing time, and are assigned to the solvent-exposed His87 ligand. The approximately 10 cm(-1) differences between His37 and His87 are attributed to the effect of H-bonding on the imidazole ring modes. The UVRR spectra of the Cu(I) protein in (2)H(2)O reveal a 1408 cm(-1) band, characteristic of NH-(2)H-exchanged histidinium, which grows in as the p(2)H is lowered. Its intensity follows a titration curve with pK(a)=4.6. This protonation is assigned to the His87 residue, whose bond to the Cu(I) is known from crystallography to be broken at low pH. As the 1408 cm(-1) band grows, a band at 1345 cm(-1) diminishes, while another, at 1337 cm(-1) stays constant. These are assigned to modes of bound His87 and His37, respectively, shifted down 7-9 cm(-1) from their Cu(II) positions.
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Base de dados:
MEDLINE
Assunto principal:
Plastocianina
/
Cobre
/
Histidina
Idioma:
En
Ano de publicação:
2002
Tipo de documento:
Article