Guanidinium chlorid

Rideout and Breslow first reported [2a] the kinetic data for the accelerating effect of water, for the Diels Alder reactions of cyclopentadiene with methyl vinyl ketone and acrylonitrile and the cycloaddition of anthracene-9-carbinol with N-ethylmaleimide, giving impetus to research in this area (Table 6.1). The reaction in water is 28 to 740 times faster than in the apolar hydrocarbon isooctane. By adding lithium chloride (salting-out agent) the reaction rate increases 2.5 times further, while the presence of guanidinium chloride decreases it. The authors suggested that this exceptional effect of water is the result of a combination of two factors the polarity of the medium and the... 

Investigation of the polypeptide folding properties of iron-sulfur via NMR spectroscopy began in 1997 (122). All studies performed to date have focused on the effect of the addition of guanidinium chloride (GdmCl hereafter) to protein solutions. Under these conditions, re-... 

Many examples of the phase-transfer catalysed epoxidation of a,(3-unsaturated carbonyl compounds using sodium hypochlorite have been reported [e.g. 7-10]. The addition of transition metal complexes also aids the reaction [11], but advantages in reaction time or yields are relatively insignificant, whereas the use of hexaethyl-guanidinium chloride, instead of a tetra-alkylammonium salt, enhances the rate of epoxidation while retaining the high yields (>95%) [10]. Intermediate (3-haloalkanols are readily converted into the oxiranes under basic conditions in the presence of benzyltriethylammonium chloride [12]. 

The extent of formation of protein disulfides with time was determined by withdrawing aliquots which were acidified to pH 5.5 and alkylated with N-ethylmaleimide. The disulfide content of the peptide was determined after its isolation. Formation of two intrapeptide disulfide bonds proceeded at the same rate (within experimental error) as formation of the first two disulfides in reduced lysozyme. The first-order rate constant for these two processes (0.5 min-1) was eight times that describing the rate of oxidation of reduced lysozyme in the presence of 6 M guanidinium chloride, suggesting substantial specificity in the process in absence of denaturant. An additional indication of specificity was the finding that 13-105 reached its maximum of two —S—S— bonds in less than 20 minutes, retaining one reduced thiol from 20 to 240 minutes. For subsequent studies this material was S-alkylated with N-ethylmaleimide. 

Fig. 4. Equilibrium curves for the unfolding and refolding of penicillinase in guanidinium chloride measured by viscosity (1), difference spectroscopy (2), and mean residue rotation [m jiM (3). Redrawn with permission from Robson and Pain (1976b).

Sola-Penna, M., A. Ferreira-Pereira, A.P. Lemos, and J.R. Meyer-Femandes. 1997. Carbohydrate protection of enzyme structure and function against guanidinium chloride treatment depends on the nature of carbohydrate and enzyme. Eur J Biochem 248 24—29. 

Very recently evidence was provided that Hmd contains a low-molecular-mass, thermolabile cofactor that is tightly bound to the enzyme but could be released upon enzyme denaturation in urea or guanidinium chloride (Buurman et al. 2000). No indications were found that the cofactor contains a redox-active transition metal. Further studies are needed to determine the structure of the cofactor and its putative role in the catalytic mechanism. 

After separation, wash the gel with two bed volumes of 2 M KCl or 6 M guanidinium chloride and re-equilibrate it. If this purification is not sufficient, cycle the gel as described in Sect. 3.4.1. 

The contribution of the poly(Pro)II conformation to the ensemble of unordered peptides has been considered.1158 The temperature dependence of [0]222 for the peptide Ac-YEAAAKEAPAKEAAAKA-NH2 in 8 M guanidinium chloride and of poly(Lys) in water and in ethylene glycol/water (2 1) mixtures 156 was fitted to a two-state equation for a poly(Pro)II-unordered equilibrium with a temperature-independent AH and temperature-independent molar ellipticities for the two components. The peptide with a Pro at the central position is an unordered peptide, the spectrum of which has pronounced poly(Pro)II-like features at low temperatures. This fit yielded [0]222=- -9580 deg-cm2dmol 1 for the poly (Pro)II component and —5560 deg-cm2-dmol 1 for the unordered component. These values provide a method for roughly estimating the poly(Pro)II content, /Pn, of an unordered peptide from [0]222 ... 

Wild-type PYP was prepared as described in [10]. Samples of PYP in native conformation were prepared in 10 mM phosphate buffer (pH = 7.5), using HPLC grade water. Denaturation of PYP was achieved by adding solid guanidinium chloride to 7 M final concentration. After denaturation, the pH was set to 9.6 with KOH, in order to deprotonate the chromophore. The absorbance of each sample was adjusted to 1 per mm at the absorption maximum. 

An extreme conformational alteration is the denaturation of proteins, which may be caused by heating or by treatment with reagents such as strong acids and bases, urea, guanidinium chloride, and sodium dodecyl sulfate (SDS). 

A series of reference proteins of known molecular masses are used to calibrate the column and Mr for an unknown protein is estimated from its position on the graph.195,196 Another modification of the method depends upon chromatography in a high concentration of the denaturing salt guanidinium chloride. The assumption is made that proteins are denatured into random coil conformations in this solvent.196... 

Polypeptide Chain Molecular Weight Determination by Gel Permeation Studies on Agarose Columns in 6M Guanidinium Chloride... 

Gel permeation chromatography of protein linear random coils in guanidinium chloride allows simultaneous resolution and molecular weight analysis of polypeptide components. Column calibration results are expressed in terms of a log M vs. Kd plot or of effective hydrodynamic radius (Re/). For linear polypeptide random coils in 6M GuHCl, Re is proportional to M0 555, and M° 555 or Re may be used interchangeably. Similarly, calibration data may be interpreted in terms of N° 555 (N is the number of amino acid residues in the polypeptide chain), probably the most appropriate calibration term provided sequence data are available for standards. Re for randomly coiled peptide heteropolymers is insensitive to amino acid residue side-chain composition, permitting incorporation of chromophoric, radioactive, and fluorescent substituents to enhance detection sensitivity. 

---