Research topic
In silico screening of putative N-acetyl-β-D-hexosaminidases, α -D-glucuronidases, and β-D-glucuronidases will be carried out in order to find out novel biocatalysts for the production of new derivatives of N-acetylglucosamine. In addition, novel lipases will be also searched for the synthesis of sorbitol- and sugar-based esters in green solvents. Likewise, we will look for microbial enzymes for the degradation of polyester-type plastics such as polyhydroxyalkanoate (PHAs), polybutylene succinate (PBS), polybutylene succinate-co-butylene adipate (PBSA), poly(ε-caprolactone) (PCL) or poly(L-lactic acid) (PLA). Although some enzymes have been described for these purposes, novel improved biocatalysts are demanded. So, exploration of new biocatalysts is necessary, and actinomycetes could be an interesting alternative as a microbial source of novel enzymes. As a starting point, some interesting gene sequences encoding these enzymes have been found in some genomes deposited by the UCM Enzyme Biotechnology group in the GenBank. Several enzyme candidates will be chosen to clone the corresponding synthetic genes into different plasmids (such as pET28 or pET24), and employing different expression hosts (E. coli, Rhodococcus or Streptomyces). Fermentative production and further purification of the enzymes from the culture broths of the recombinant strains will be performed using different purification techniques. Protein structure and kinetic features of purified enzymes will be studied. Different tailored-made biocatalysts will be created by protein engineering techniques (such as directed evolution, site-directed mutagenesis, etc.). If necessary, immobilization will be carried out to improve the robustness and allow multiple use of the biocatalysts.