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Quaternary pyrochlore-type solid solutions, CaGdZrNb(1-x)Ta(x)O(7) (x = 0, 0.2, 0.4, 0.6, 0.8, 1), were prepared by a high-temperature ceramic route. The pyrochlore phases of the compounds were confirmed by powder X-ray diffraction (XRD), Raman spectroscopy, and transmission electron microscopy. The crystallographic parameters of the pyrochlore compounds were accurately determined by Rietveld analysis of the powder XRD data. The isovalent substitution of Ta in place of Nb at the B site can reveal the effect of chemical bonding on lattice thermal expansion and oxide ion conductivity because both Nb and Ta have the same ionic radius (0.64 Å). Lattice thermal expansion coefficients of the samples were calculated from high-temperature XRD measurements, and it was found that the thermal expansion coefficient decreases with substitution of Ta. Oxide ion conductivity measured by a two-probe method also shows the same trend with substitution of Ta, and this can be attributed to the high bond strength of the Ta-O bond compared to that of the Nb-O bond. Microstructural characterization using scanning electron microscopy proves that the size of the grains has a small effect on the oxide ion conductivity. Our studies established the role of chemical bonding in deciding the conductivity of pyrochlore oxides and confirmed that the 48f-48f mechanism of oxide ion conduction is dominant in pyrochlore oxides.

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The effect of simultaneous substitutions of Ca at A site and Nb or Ta at B site in pyrochlore-type solid solutions: (Ca(x)Gd(1-x))(2)(Zr(1-x)M(x))(2)O(7) (x = 0.1, 0.2, 0.3, 0.4, 0.5 and M = Nb or Ta) were studied by powder X-ray diffraction (XRD), FT NIR Raman spectroscopic techniques and transmission electron microscopy. The solid solutions were prepared by the conventional high-temperature ceramic route. The XRD results and Rietveld analysis revealed that the defect fluorite structure of Gd(2)Zr(2)O(7) progressively changed to a more ordered pyrochlore phase by simultaneous substitutions at A and B sites. Raman spectroscopy reveals the progressive ordering in the anion sublattice with simultaneous doping. High-resolution images and selected-area electron diffraction patterns obtained from TEM confirms the XRD and Raman spectroscopic results. High-temperature XRD studies show that the lattice expansion coefficient in these pyrochlore oxides is of the order of 10(-6) K(-1). Lattice thermal expansion coefficient increases with increase of disorder in pyrochlore oxides, and hence the variation of thermal expansion coefficient with composition is also a good indicator of disordering in pyrochlore-type oxides. The ionic conducting properties of the samples were characterised by impedance spectroscopy, and it was found that Nb-doped compositions show a considerable change in conductivity near the phase boundary of disordered pyrochlore and defect fluorite phases.

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Serine hydroxymethyltransferase from mammalian and bacterial sources is a pyridoxal-5'-phosphate-containing enzyme, but the requirement of pyridoxal-5'-phosphate for the activity of the enzyme from plant sources is not clear. The specific activity of serine hydroxymethyltransferase isolated from mung bean (Vigna radiata) seedlings in the presence and absence of pyridoxal-5'-phosphate was comparable at every step of the purification procedure. The mung bean enzyme did not show the characteristic visible absorbance spectrum of a pyridoxal-5'-phosphate protein. Unlike the enzymes from sheep, monkey, and human liver, which were converted to the apoenzyme upon treatment with l-cysteine and dialysis, the mung bean enzyme similarly treated was fully active. Additional evidence in support of the suggestion that pyridoxal-5'-phosphate may not be required for the mung bean enzyme was the observation that pencillamine, a well-known inhibitor of pyridoxal-5'-phosphate enzymes, did not perturb the enzyme spectrum or inhibit the activity of mung bean serine hydroxymethyltransferase. The sheep liver enzyme upon interaction with O-amino-d-serine gave a fluorescence spectrum with an emission maximum at 455 nm when excited at 360 nm. A 100-fold higher concentration of mung bean enzyme-O-amino-d-serine complex did not yield a fluorescence spectrum. The following observations suggest that pyridoxal-5'-phosphate normally present as a coenzyme in serine hydroxymethyltransferase was probably replaced in mung bean serine hydroxymethyltransferase by a covalently bound carbonyl group: (a) inhibition by phenylhydrazine and hydroxylamine, which could not be reversed by dialysis and or addition of pyridoxal-5' phosphate; (b) irreversible inactivation by sodium borohydride; (c) a spectrum characteristic of a phenylhydrazone upon interaction with phenylhydrazine; and (d) the covalent labeling of the enzyme with substrate/product serine and glycine upon reduction with sodium borohydride. These results indicate that in mung bean serine hydroxymethyltransferase, a covalently bound carbonyl group has probably replaced the pyridoxal-5'-phosphate that is present in the mammalian and bacterial enzymes.

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The interaction of aminooxy compounds such as aminooxyacetate (AAA), L-canaline, and hydroxylamine with sheep liver serine hydroxymethyltransferase (EC 2.1.2.1) was studied by absorption spectra and stopped-flow spectrophotometry and compared with the unique feature of interaction of O-amino-D-serine (OADS) with the enzyme [Baskaran, N., Prakash, V., Appu Rao, A. G., Radhakrishnan, A. N., Savithri, H. S., & Appaji Rao, N. (1989) Biochemistry (preceding paper in this issue)]. The reaction of AAA (0.5 mM) with the Schiff base of the enzyme resulted in the formation of pyridoxal 5'-phosphate (PLP) and was biphasic with rate constants of 191 and 19 s-1. The formation of the PLP-AAA oxime measured by decrease in absorbance at 388 nm on interaction of AAA with the enzyme had a rate constant of 5.2 M-1 s-1. On the other hand, the reaction of L-canaline with the enzyme was slower as measured by the disruption of enzyme-Schiff base than the reaction of OADS and AAA. In contrast, the formation of PLP as an intermediate could not be detected upon the interaction of hydroxylamine with the enzyme. The reaction of D-cycloserine with the enzyme was much slower (1.6 x 10(2) M-1 s-1) than the aminooxy compounds. These observations indicate that the aminooxy compounds that are structural analogues of serine (OADS, AAA, and canaline) formed PLP as an intermediate prior to the formation of oxime, whereas with hydroxylamine such an intermediate could not be detected.

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The mechanism of interaction of O-amino-D-serine (OADS) with sheep liver serine hydroxymethyltransferase (EC 2.1.2.1) (SHMT) was established by measuring changes in the enzyme activity, absorption spectra, circular dichroism (CD) spectra, and stopped-flow spectrophotometry. OADS was a reversible noncompetitive inhibitor (Ki = 1.8 microM) when serine was the varied substrate. The first step in the interaction of OADS with the enzyme was the disruption of enzyme-Schiff base, characterized by the rapid disappearance of absorbance at 425 nm (6.5 X 10(3) M-1 s-1) and CD intensity at 430 nm. Concomitantly, there was a rapid increase in absorbance and CD intensity at 390 nm. The spectral properties of this intermediate enabled its identification as pyridoxal 5'-phosphate (PLP). These changes were followed by a slow unimolecular step (2 X 10(-3) s-1) leading to the formation of PLP-OADS oxime, which was confirmed by its absorbance and fluorescence spectra and retention time on high-performance liquid chromatography. The PLP-OADS oxime was displaced from the enzyme by the addition of PLP as evidenced by the restoration of complete enzyme activity as well as by the spectral properties. The unique feature of the mechanism proposed for the interaction of OADS with sheep liver SHMT was the formation of PLP as an intermediate.

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Rabbit intestinal glucoamylase-maltase was examined in detail with respect to its molecular weight, sedimentation, diffusion and viscosity. It is a large asymmetrical molecule, with a molecular weight of 750 000-760 000. Its appearance under the electron microscope supports the idea that it is a long string (62.0 nm) consisting of eight beads of diameter 6.0 nm each and a surface-to-surface interbead distance of approx. 2.0 nm. The shape of the enzyme derived from its hydrodynamic behaviour by using the string-of-spherical-beads model originally proposed by Kuhn [(1932) Z. Phys. Chem. Abt. A 161, 1-32] and later modified by Shulman [(1953) J. Am. Chem. Soc. 75, 5846-5852] fits moderately well with the electron-microscopic picture. The beads might represent about six subunits, and the absence of sulphur from the enzyme and the inability to dissociate the enzyme by conventional methods indicate the possibility of unusual covalent cross-linking between the subunits and between the beads.

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1. Intestinal absorption of glycylglycine was studied in four control subjects and six patients with tropical sprue by using a direct technique of intestinal perfusion. 2. The patients with tropical sprue showed significant impairment in the absorption of the dipeptide.

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The effect of harmaline on amino acid and dipeptide uptake was studied in the monkey. Harmaline inhibited both the systems. Preincubation of the tissue with 4 mmole/1 harmaline for 10 min was necessary for maximal inhibition. The uptake of glycyl-l-leucine- ((14)C) and ((14)C)-glycyl-l-leucine was inhibited to the same extent. The uptake of glycyl-l-proline, a dipeptide which was presumably taken up intact by the mucosal cells, was also inhibited. Total replacement of sodium in the medium reduced the uptake of both glycyl-l-leucine and glycyl-l-proline. But the uptake of these dipeptides was not inhibited by harmaline in the absence of sodium. However, the sodium-independent uptake of glycyl-l-leucine and glycyl-l-proline was also inhibited by other dipeptides.

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1. There are two saturable transport processes in the monkey small intestine for glycyl-L-leucine, one with Vmax. 1 mumol min-1 g-1 wet weight of tissue and an affinity constant (kt) of 5 mmol/l, and the other with Vmax 3.9 mumol min-1 g-1 wet weight of tissue and kt 33 mmol/l. 2. Glycyl-L-leucine uptake is inhibited by a wide variety of amino acids, although to a variable extent. The inhibition was shown to be competitive with leucine used as the representative amino acid. Phenylalanine, methionine, alanine and leucine are the most potent in their inhibitory action. 3. The effect of various amino acids on the hydrolysis of glycyl-L-leucine by particulate and cytosol fractions of monkey small intestine was studied. All the amino acids, except glycine, proline, alanine and glutamic acid, inhibit both the particulate and cytosol glycyl-L-leucine hydrolase activities. In general, the cytosol enzyme is more susceptible to amino acid inhibition than the particulate enzyme. 4. There is no correlation between the effects of amino acids on glycyl-L-leucine uptake and hydrolysis of glycyl-L-leucine by either particulate or cytosol fraction.

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Three untreated phenylketonuric Indian children aged respectively 3 1/2 years, 1 1/2 years and 1 year showed rapid neurological deterioration. Plasma, cerebrospinal fluid and urine phenylalanine concentrations were significantly raised and the phenylalanine-tyrosine ratio was high. Analysis of a biopsy of the right frontal lobe of the brain in one case showed the myeline lipids--cerebroside and sulphatide--to be decreased. The total cerebroside in white matter was low. Light microscopy showed marked pallor of the white matter of the brain and extensive spongy degeneration. Ultrastructurally these spongy vesicles are located between the lamellae of the myelin sheath.

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Among the powerful dipeptidases of the cytosol fraction the well-characterized 'master' dipeptidase of broad substrate specificity is an example of a 'true' dipeptidase. There are only a limited number of peptidases which together can hydrolyse the total range of the theoretically possible dipeptides. In the lumen, the dipeptides of dietary origin apparently enter the cell through a single transport system of broad specificity in the monkey and possibly also in man. Perfusion studies indicated impared absorption of glycine and Gly-Gly in Indian compared to English subjects, but the kinetic advantage of dipeptide uptake over that of the free amino acid was maintained. In patients with tropical sprue, apart from decreased peptide absorption, there was an increased 'backflow' of the constituent amino acids. The back-flow assumes nutritional significance since dipeptide uptake is competitively inhibited by amino acids of a certain specificity. Inhibition of brush border glycylglycine hydrolase (glycylglycine dipeptidase, EC 3.4.13.1) by L- leucine is non-competitive while that of cytosol enzyme is competitive. Certain other amino acids have no effect on glycylleucine hydrolase (glycylleucine dipeptidase, EC 3.4.13.2) from either fraction but inhibit Gly-Leu uptake. Thus, the inhibition of dipeptide uptake appears to be a consequence of interaction of amino acids directly with the dipeptide transport system and not with dipeptidases.

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Intestinal absorption of glycine 20 mmol/1, glycyl-glycine 10 mmol/1 plus L-leucine 10 mmol/1, and glycyl-L-leucine 10 mmol/1 has been studied by intestinal perfusion in 11 patients with tropical sprue and 10 control subjects. The patients with sprue had a significant reduction in the rate of absorption of glycine from a 20 mmol/1 solution, but there were no significant differences in the absorption of the other substances. The failure to demonstrate any difference in the absorption of these substances is probably related to their low concentration relative to the maximum absorptive capacity of the intestine. In both groups of subjects the kinetic advantage of glycyl-glycine absorption as compared with glycine absorption was maintained. When the dipeptides were perfused, free amino acids appeared in the perfusate presumably by "back diffusion" from the mucosal cells. In the case of glycyl-L-leucine considerably more glycine and leucine were found in the perfusate in patients with sprue than in the control subjects. There was no correlation between peptide absorption and the concentration of total glycly-glycine hydrolase and glycyl-L-leucine hydrolase, measured as combined brush border and cytosol enzymes. The concentrations of these enzymes were similar in both groups of subjects.

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