Histidine separation

In recent studies employing NMR it was possible to identify signals from the C2 protons of each histidine separately (see Table XIV). The thermodynamic parameters are also provided for each histidine in... 

Crystallization Method. Such methods as mechanical separation, preferential crystallisation, and substitution crystallisation procedures are included in this category. The preferential crystallisation method is the most popular. The general procedure is to inoculate a saturated solution of the racemic mixture with a seed of the desired enantiomer. Resolutions by this method have been reported for histidine (43), glutamic acid (44), DOPA (45), threonine (46), A/-acetyl phenylalanine (47), and others. In the case of glutamic acid, the method had been used for industrial manufacture (48). 

The crude product is dissolved in five times its weight of water, and after clearing with a little Norite the solution is diluted with one and one-half volumes of 95 per cent alcohol. The product separates in well-formed, snow-white crystals, and after standing for several days in an ice chest is collected with suction on a Buchner funnel. The yield of purified histidine monohydrochloride is 75-80 g. (Note 5). The compound melts at 251-2520, with decomposition. The amino acid is not race-mized by the procedure employed, and shows the characteristic optical activity, [a]n6° = +8.00, in the presence of three moles of... 

Figure 39-14. Zinc fingers are a series of repeated domains (two to nine) in which each is centered on a tetrahedral coordination with zinc. In the case of TFIIIA, the coordination is provided by a pair of cysteine residues (C) separated by 12-13 amino acids from a pair of histidine (H) residues. In other zinc finger proteins, the second pair also consists of C residues. Zinc fingers bind in the major groove, with adjacent fingers making contact with 5 bp along the same face of the helix.

Flgvire 4.5 The influence of endcapping on peak shape and retention of soee PTH-anino acids using a reversed-phase separation system. Peak identification 1 PTH-histidine, 2 PTH-arginine and 3 PTH-valine. (Reproduced with permission from ref. 71. Copyright Preston Publications, Inc.)... 

Figure 8.43 Separation of enantiomers using complexation chromatography. A, Separation of alkyloxiranes on a 42 m x 0.2S mm I.O. open tubular column coated with 0.06 M Mn(II) bis-3-(pentafluoro-propionyl)-IR-camphorate in OV-ioi at 40 C. B, Separation of D,L-amino acids by reversed-phase liquid chromatography using a mobile phase containing 0.005 M L-histidine methyl ester and 0.0025 M copper sulfate in an ammonium acetate buffer at pH 5.5. A stepwise gradient using increasing amounts of acetonitrile was used for this separation.

The dinuclear active site of urease (1) has been studied in great detail23-29 and has inspired manifold model studies—hence a separate section, Section 6.3.4.12.7, is dedicated to the coordination chemistry related to urease. E. coli Glx I is the first example of a Ni-dependent isomerase and contains a single Ni11 ion coordinated by two histidines, two axial carboxylates of glutamic acid, and two water molecules (2).30-32 It is not active with Zn bound, which is believed to result from the inability of the Zn-substituted enzyme to bind a second aqua ligand and to adopt a six-coordinate structure. 

Valproic acid has been determined in human serum using capillary electrophoresis and indirect laser induced fluorescence detection [26], The extract is injected at 75 mbar for 0.05 min onto a capillary column (74.4 cm x 50 pm i.d., effective length 56.2 cm). The optimized buffer 2.5 mM borate/phosphate of pH 8.4 with 6 pL fluorescein to generate the background signal. Separation was carried out at 30 kV and indirect fluorescence detection was achieved at 488/529 nm. A linear calibration was found in the range 4.5 144 pg/mL (0 = 0.9947) and detection and quantitation limits were 0.9 and 3.0 pg/mL. Polonski et al. [27] described a capillary isotache-phoresis method for sodium valproate in blood. The sample was injected into a column of an EKI 02 instrument for separation. The instrument incorporated a conductimetric detector. The mobile phase was 0.01 M histidine containing 0.1% methylhydroxycellulose at pH 5.5. The detection limit was 2 pg/mL. 

Even though the iron atoms are separated in haemoglobin by about 25 A, communication between them is still able to occur and this has been postulated to involve a trigger mechanism (Perutz, 1971). The trigger is the movement of the proximal histidine as dioxygen binds to (or is released from) the Fe(n) and results in interconversion between the T and R structures. This movement causes a conformational change which is transmitted through the protein to the other iron sites. X-ray studies indicate that relative shifts of up to 6 A at subunit interfaces occur between the T and R states (Perutz, 1978). 

With multiple ionizable groups, such as in amino acids and proteins, each group titrates separately according to its pKa. The titration curves shown in Fig. 23-5 are for the amino acids glycine, histidine, and glutamate. 

In the deoxygenated form, each Cu(I) centre is coordinated to three nitrogen atoms of three histidines. The great Cu/Cu separation (4.62 A) foreshadows the presence of a cavity able to host a dioxygen molecule. In fact, in the oxygenated form, the oxygen molecule is planarily positioned according to a p-rj2 r]2 peroxo coordination, and the Cu/Cu separation decreases (—3.5 A). 

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