Orientation of Cutinase Adsorbed onto PMMA Nanoparticles Probed by Tryptophan Fluorescence
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- 作者：Andrea M. Santos ; Aleksander Fedorov ; José ; M. G. Martinho ; Ricardo P. Baptista ; Maria  ; ngela Taipa ; Joaquim M. S. Cabral
- 刊名：Journal of Physical Chemistry B
- 出版年：2008
- 出版时间：March 27, 2008
- 年：2008
- 卷：112
- 期：12
- 页码：3581 - 3585
- 全文大小：143K
- 年卷期：v.112,no.12(March 27, 2008)
- ISSN：1520-5207
文摘
The fluorescence of the single tryptophan (Trp69) of cutinase from Fusarium solani pisi, free in aqueoussolution and adsorbed onto the surface of poly(methyl methacrylate) (PMMA) latex particles, was studied atpHs of 4.5 and 8.0. The monodisperse PMMA particles (d = 106.0 ± 0.1 nm) were coated with a quitecompact monolayer of cutinase at both pH values. The Trp decay curve of the folded free cutinase in solutioncan only be fitted with a sum of four exponentials with lifetimes of 0.05, 0.3-0.4, 2-3, and 6-7 ns, irrespectiveof pH. The 50 ps lifetime is attributed to the population of Trp residues hydrogen bonded with the Ala32 andstrongly quenched by a close disulfide bridge, while the other lifetimes are due to the non-hydrogen-bondedTrp rotamers. The 50 ps Trp lifetime component disappears by temperature melting and upon protein adsorption,owing to the disruption of the Trp-Ala hydrogen bond and the release of the Trp residue from the vicinityof the disulfide bridge. This shows that cutinase adsorption occurs by the region of the protein where the Trpis located, which agrees with the retention of cutinase enzymatic activity by adsorption at basic pH.