Quenching of beta-lactoglobulin fluorescence by 2-nitro-5-thiobenzoic acid.
نویسندگان
چکیده
I3C-NMR studies of modified 6-lactoglobulins, cyanylated at the single thiol group, have provided evidence to support the location of the thiol at Cys-121 [ I J and useful information about the nuclear relaxation behaviour of the thiocyanate carbons of cyanylated thiol groups in proteins [2]. The O-lactoglobulin monomer contains two tryptophan residues. Trp-19 is buried deep inside the hydrophobic 6-barrel core of the protein and Trp61 is located in a flexible loop. The thiol group of Cys-121 is located in a hydrophobic cleft near the surface of the protein [3]. The binding of retinol by 6-lactoglobulin significantly quenches the inbinsic fluorescence of the protein [4]. Trp-19 is the only invariant residue in the lipocalin superfamily, and it has been implicated in the stabilisation of retinol at it's binding site, although it is not essential for retinol binding [51. The intrinsic fluorescence of 6-lactoglobulin is mainly due to Trp-19, since the fluorescence of a W19A mutant protein is only 20 % of that of wild-type protein [6]. The mixed disulphide derivative of 0-lactoglobulin B with 2-nitro-5-thiobenzoic acid (TNB) was prepared using 5,5'dithiobis(2,T-nitrobenzoic acid) (DTNB) as previously described 111. Fluorescence measurements were made at 25 O C using a Hitachi Model F-4500 fluorimeter. Fluorescence emission spectra of 8-lactoglobulin B and the mixed disulphide derivative were recorded using an excitation wavelength of 290 nm and excitation and emission slit widths were fixed at 4 nm. 1 ml 8lactoglobulin solutions used for fluonmetry contained 6-10 pM monomer, in 10 mM potassium phosphate buffer at pH 7.0. Distances between amino acid side-chains in 6-lactoglobulin were measured using SwissPdbViewer version 2.1 from the Glaxo Institute for Molecular Biology, Geneva, Switzerland. Coordinates for bovine 0-lactoglobulin were kindly supplied by L. Sawyer, Structural Biochemistry Group, University of Edinburgh. The intrinsic tryptophan fluorescence of 8-lactoglobulin B has a maximum emission wavelength at 335 nm (Fig. IA), indicative of a hydrophobic environment around the tryptophan residues.
منابع مشابه
Aerobic oxidation of p-hydroquinone by horse radish peroxidase in the presence of a thiol and MnCl2.
In the presence of MnCl2 and a thiol (glutathione, cysteine, 2-nitro-5-thiobenzoic acid) horse radish peroxidase oxidizes p-hydroquinone to p-benzoquinone which in turn immediately adds the thiol present yielding 2-S-substituted p-hydroquinone.
متن کاملThe Quenching of Pyrochlorophyll Fluorescence by Nitro Compounds
We have examined the quenching of pyrochlorophyll fluorescence by a series of substituted nitrobenzenes in ethanol-pyridine solution and the effect of fluorescence quenching on the quantum yield of sensitized photoreduction of three of the nitro compounds by hydrazobenzene. All nitro compounds examined are at least moderately good quenchers of fluorescence. There is a good correlation between t...
متن کاملInteractions of β-lactoglobulin with Cationic Surfactants: Spectroscopy Study
The interactions of β-lactoglobulin AB in the presence of cationic surfactants such as Cetyltrimethylammonium bromide and Cetyltrimethylammonium p-Toluenesulfonate have been investigated using a variety of experimental techniques such as conductivity, UV-Vis spectrophotometry and fluorimetry. The conductivity of surfactants aqueous solutions with β-lactoglobulin shows that the cmc of cationic s...
متن کاملThe fluorescence Quenching Study of Quinine in Presence of Some Anions
The quenching of quinine fluorescence intensity in the presence of some anions in aqueous solution at ambient temperature has been investigated. The quenching is found to be collisional or dynamical in nature. This study reveals the order of two groups of quencher: NaI > NaBr > NaCl > NaF and K2Cr2O7 > KMnO4 >Na2SO4 > NaClO3. Increasing anion size in the both groups leads to an increase in the...
متن کاملMCR of the quenching of the EEM of fluorescence of Aflatoxins (B1, G1) by Gold nanoparticles
In This research, gold nanoparticles were synthesized and functionalized by the antibody of aflatoxins. The quenching of the fluorescence of excitation emission matrices (EEM) of two type of aflatoxins (B1, G1), provoked by the gold nanoparticles, was studied by principal component analysis (PCA) and multivariate curve resolution with alternating least squares (MCR-ALS). These aflatoxins show q...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Biochemical Society transactions
دوره 26 1 شماره
صفحات -
تاریخ انتشار 1998