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8 páginas, 5 figuras, 3 tablas -- PAGS nros. 1099-1106, An extracellular sterol esterase from Ophiostoma piceae efficiently hydrolyzes sterol esters, triglycerides and p-nitrophenol esters. cDNA was screened with a probe obtained by PCR using as primers oligonucleotides corresponding to the N-terminal and internal mature enzyme sequences and complete sequence was obtained by 3′ rapid amplification of cDNA end (RACE) and inverse PCR. The O. piceae esterase gene had a length of 1.8 kbp and lacked introns. A search for proteins with related amino acid sequences revealed around 40% identity with lipases from Candida rugosa and Geotrichum candidum. Modelling the O. piceae enzyme, using the crystal structures of Lip1 and Lip3 from C. rugosa as templates, revealed a similar substrate-binding site, but some changes affecting the flap zone and the aromatic region of the tunnel may be responsible for the wide substrate specificity of this interesting sterol esterase. The ability of the new fungal esterase to hydrolyze triglycerides and esters of p-nitrophenol and cholesterol was compared with those of commercial lipases and cholesterol esterases showing the new enzyme the highest efficiency hydrolyzing triglycerides and sterol esters in the conditions assayed (in presence of Genapol X-100). Finally, the O. piceae gene was successfully expressed in Pichia pastoris, as a model organism to express fungal enzymes, resulting in higher levels of esterase activity than those obtained in the O. piceae cultures. In spite of its higher glycosylation degree, the recombinant enzyme was able to hydrolyze more efficiently than native enzyme the assayed substrates, This study has been supported by the Spanish projects BIO2003-00621, BIO2006- 11393 and S-0505/AMB0100. V. Barba acknowledges the financial support received from the MICINN, Spain

The chemical analysis of the acetone, chloroform, toluene and methanol extracts of a pitch sample was carried out by IR and GC-MS, leading to the identification of sixty nine compounds, including fatty acids, alcohols and hydrocarbons. Analysis of the acetone extractive of a eucalyptus wood used in Brazil for pulp production was also carried out, resulting in identification of fifty nine compounds, including mainly fatty acids, phenolic compounds, beta-sitosterol and other steroids. This analysis showed that pitch formation had a contribution from wood extractives and other sources of contamination. The results obtained and the methodology applied can be used by the pulp industry to develop new methods of pitch control.