Immobilization of recombinant human endostatin to different supports: potential applications in biomedicine

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Characterization of a novel subgroup of extracellular medium-chain-length polyhydroxyalkanoate depolymerases from Actinobacteria

Nineteen medium-chain-length (mcl) poly(3-hydroxyalkanoate) (PHA)-degrading microorganisms were isolated from natural sources. From them, seven Gram-positive and three Gram -negative bacteria were identified. The ability of these microorganisms to hydrolyze other biodegradable plastics, such as short-chain-length (scl) PHA, poly(ε-caprolactone) (PCL), poly(ethylene succinate) (PES), and poly(L-lactide) (PLA), has been studied. On the basis of the great ability to degrade different polyesters, Streptomyces roseolus SL3 was selected, and its extracellular depolymerase was biochemically characterized. The enzyme consisted of one polypeptide chain of 28 kDa with a pI value of 5.2. Its maximum activity was observed at pH 9.5 with chromogenic substrates. The purified enzyme hydrolyzed mcl PHA and PCL but not scl PHA, PES, and PLA. Moreover, the mcl PHA depolymerase can hydrolyze various substrates for esterases, such as tributyrin and p-nitrophenyl (pNP)-alkanoates, with its maximum activity being measured with pNP-octanoate. Interestingly, when poly(3-hydroxyoctanoate-co-3-hydroxyhexanoate [11%]) was used as the substrate, the main hydrolysis product was the monomer (R)-3-hydroxyoctanoate. In addition, the genes of several Actinobacteria strains, including S. roseolus SL3, were identified on the basis of the peptide de novo sequencing of the Streptomyces venezuelae SO1 mcl PHA depolymerase by tandem mass spectrometry. These enzymes did not show significant similarity to mcl PHA depolymerases characterized previously. Our results suggest that these distinct enzymes might represent a new subgroup of mcl PHA depolymerases.This work was carried out in the framework of the IP project Sustainable Microbial and Biocatalytic Production of Advanced Functional Materials (BIOPRODUCTION/NMP-2-CT-2007-026515), funded by the European Commission and by the Spanish Ministry of Education and Science (BIO2007-28707-E and CTQ2011-25052), and UPV/EHU (GIU07/55 and GIU11/25). M.S. and J.G. were the recipients of scholarships from the Spanish Ministry of Education. P(3HO) was kindly supplied by Biopolis, S.A. (Valencia, Spain), and CPI (Newcastle, United Kingdom). P(3HP) was kindly donated by CIBA (Manchester, United Kingdom).Peer Reviewe