The performance of fungal xylan-degrading enzyme preparations in elemental chlorine-free bleaching for Eucalyptus pulp

Cellulase-free xylan-degrading enzyme preparations from Acrophialophora nainiana, Humicola grisea var. thermoidea and two Trichoderma harzianum strains were used as bleaching agents for Eucalyptus kraft pulp, prior to a chlorine dioxide and alkaline bleaching sequence. In comparison to the control sequence (performed without xylanase pretreatment), the sequence incorporating enzyme treatment was more effective. Removal of residual lignin was indicated by a reduction in kappa number. Overall, enzyme preparations from T. harzianum were marginally more effective in reducing pulp viscosity and chlorine chemical consumption and improving the brightness of the kraft pulp. However, the highest reduction in pulp viscosity was mediated by the xylanase preparation from A. nainiana. Xylanase pretreatment compares very favorably with that of chemical pulping. Journal of Industrial Microbiology & Biotechnology (2002) 28, 204–206 DOI: 10.1038/sj/jim/7000227 Received 27 April 2001/ Accepted in revised form 03 November 2001     

Microsatellite evolution--a reciprocal study of repeat lengths at homologous loci in cattle and sheep

The application of microsatellites in evolutionary studies requires an understanding of the patterns governing their evolution in different species. The finding that homologous microsatellite loci are longer, i.e., containing more repeat units, in human and in other primates has been taken as evidence for directional microsatellite evolution and for a difference in the rate of evolution between species. However, it has been argued that this finding is an inevitable consequence of biased selection of longer-than-average microsatellites in human, because cloning procedures are adopted to generate polymorphic and, hence, long markers. As a test of this hypothesis, we conducted a reciprocal comparison of the lengths of microsatellite loci in cattle and sheep using markers derived from the bovine genome as well as the ovine genome. In both cases, amplification products were longer in the focal species, and loci were also more polymorphic in the species from which they were originally cloned. The crossing pattern that we found suggests that interspecific length differences detected at homologous microsatellite loci are the result of biased selection of loci associated with cloning procedures. Hence, comparisons of microsatellite evolution between species are flawed unless they are based on reciprocal analyses or on genuinely random selection of loci with respect to repeat length.      

Irreversible inhibition of putrescine-stimulated S-adenosyl-L-methionine decarboxylase by berenil and pentamidine.

The putrescine-stimulated S-adenosyl-L-methionine decarboxylases from rat liver and yeast were strongly inhibited by Berenil and to a lesser extent by Pentamidine. Ten times greater drug concentrations were needed to achieve a similar level of inhibition of a Mg2+-stimulated bacterial enzyme. The inhibition was irreversible in that extensive dialyses or precipitation with (NH4)2SO4 did not restore enzyme activity. Putrescine did not protect the enzyme against Berenil, but adenosylmethionine either alone or with putrescine partially protected the irreversible action of Berenil. The compound 4,4'-diamidinodiphenylamine, which differs from Berenil only in lacking the azo group between benzene rings, was a weaker inhibitor than Berenil, and its inhibition was reversible. Berenil also inhibited the activity of adenosylmethionine decarboxylase in vivo, by depressing the activity of the enzyme in normal rat liver, for at least 24 h after a single injection (50 mg/kg body wt.) of the drug.     

Stabilization of ornithine decarboxylase and N1-spermidine acetyltransferase in rat liver by methylglyoxal bis(guanylhydrazone)

The activities of ornithine decarboxylase and spermidine N1-acetyltransferase started to rise in normal rat liver 4 h after the intraperitoneal injection of methylglyoxal bis(guanylhydrazone) (MGBG; 80 mg/kg). Ornithine decarboxylase had its greatest activity 24 h after a single injection of MGBG and the acetyltransferase peaked 8 h after the injection. Measurement of the apparent half-life of ornithine decarboxylase after MGBG treatment revealed a clear decrease in the decay rate of the enzyme in both normal and regenerating rat liver. MGBG slowed the decay of the transferase also in normal rat liver, as well as inhibiting its activity in vitro. The stabilization by MGBG of these two short-lived proteins involved in metabolism of polyamines should lead to their accumulation in liver, thus explaining their increased activities. In the case of ornithine decarboxylase, studies with a specific antibody against mouse kidney ornithine decarboxylase showed that the rise in ornithine decarboxylase activity after MGBG application was not due to the appearance of an immunologically different isozyme.     

Specific inactivation of prolyl 4-hydroxylase and inhibition of collagen synthesis by oxaproline-containing peptides in cultured human skin fibroblasts

The crucial role of collagen in fibrotic disorders has prompted attempts to develop drugs that inhibit collagen accumulation. Peptides containing the unphysiological amino acid 5-oxaproline (Opr) have recently been found to act as specific syncatalytic inactivators of pure prolyl 4-hydroxylase, the enzyme that catalyzes the formation of 4-hydroxyproline in collagens. The present study indicates that oxaproline-containing peptides benzyloxycarbonyl-Phe-Opr-Gly-benzyl ester (I) and benzyloxycarbonyl-Phe-Opr-Gly-ethyl ester (II) inactivate prolyl 4-hydroxylase in cultured human skin fibroblasts, peptide I being about twice as potent as peptide II. Inactivation by 50% was observed after culturing with about 20-40 microM concentrations of peptide I for 48 h. The inactivation appears to be specific, as no changes were found in the activities of two other intracellular enzymes of collagen synthesis, lysyl hydroxylase and galactosylhydroxylysyl glucosyltransferase. Synthesis of 4-hydroxyproline by the cells was markedly decreased, and 4-hydroxyproline-deficient procollagen accumulated intracellularly, whereas no changes were found in the incorporation of [14C]leucine into protein after culturing of the cells with a 30 microM concentration of peptide I for 48 h. No changes were seen in the viability of the cells or the release of lactate dehydrogenase from them into the culture medium. No significant changes were found in the steady-state levels of the mRNAs for the pro-alpha 1 chains of type I and type III procollagens or for the alpha and beta subunits of prolyl 4-hyroxylase or fibronectin after culturing with 75 microM peptide I for 48 h. The data indicate that inactivation of cellular prolyl 4-hydroxylase has marked effects on cellular 4-hydroxyproline formation and collagen secretion but no effects on the steady-state levels of mRNAs for type I and III procollagens or the two types of subunit of prolyl 4-hydroxylase.     

Length variation in the internal transcribed spacers of ribosomal DNA in Picea abies and related species

The structure and variation of nuclear ribosomal DNA (rDNA) units of Picea abies, (L.) Karst. was studied by restriction mapping and Southern hybridization. Conspicuous length variation was found in the internal transcribed spacer (ITS) region of P. abies, although the length of this region is highly conserved both within and among most of the plant species. Two types of ITS variants (A and B), displaying a size difference of 0.5 kb in the ITS2 region, were present within individuals of P. abies from Sweden, Central Europe and Siberia. A preliminary survey of 14 additional Eurasian and North American species of Picea suggested that length variation in the ITS region is widespread in this genus. Alltogether three length variants (A, B and C) were identified. Within individuals of eight Picea species, two length variants were present within the genome (combinations of A and B variants in P. glehnii, P. maximowiczii, P. omorika, P. polita and P. sitchensis and variants B and C in P. jezoensis, P. likiangensis and P. spinulosa). Within individuals from five species, however only one rDNA variant was present in their genome (variant A in P. aurantiaca, P. engelmannii, P. glauca, P. koraiensis and P. koyamai; variant B in P. bicolor). The ITS length variation will be useful as a molecular marker in evolutionary studies of the Picea species complex, whose phylogeny is controversial. The presence of intraindividual variation in, and shared polymorphism of the, ITS length variants raises questions about the occurrence of interspecific hybridization during the evolutionary history of Picea.     

1-Aminooxy-3-aminopropane, a new and potent inhibitor of polyamine biosynthesis that inhibits ornithine decarboxylase, adenosylmethionine decarboxylase and spermidine synthase

1-Aminooxy-3-aminopropane was shown to be a potent competitive inhibitor (Ki = 3.2 nM) of homogenous mouse kidney ornithine decarboxylase, a potent irreversible inhibitor (Ki = 50 microM) of homogeneous liver adenosylmethionine decarboxylase and a potent competitive (Ki = 2.3 microM) of homogeneous bovine brain spermidine synthase. It did not inhibit homogeneous bovine brain spermine synthase and it did not serve as a substrate for spermidine synthase. The compound did not inhibit tyrosine aminotransferase, alanine aminotransferase or aspartate aminotransferase, which are pyridoxal phosphate-containing enzymes like ornithine decarboxylase. The inactivation of adenosylmethionine decarboxylase was partially prevented by pyruvate, which is the coenzyme of adenosylmethionine decarboxylase, and by the substrate, adenosylmethionine. 1-Aminooxy-3-aminopropane at 0.5 mM concentration inhibited the growth of HL-60 promyelocytic leukemia cells and this inhibition was prevented by spermidine but not by putrescine.