Target Enzyme or Product Bioprospecting for Glycohydrolase- and 1,2-Propanediol-producing Microorganisms from Geothermal Intertidal Pools in Northern Iceland Pia Maria Iloranta, Jón Pétur Jóelsson, Sean Michael Scully, Jóhann Örlygsson* *Corresponding author (jorlys@unak.is) University of Akureyri Borgir v/v Norðurslóð 600 Akureyri, Iceland Abstract Macro algae species are promising feedstocks for biotechnological applications. Unlike terrestrial lignocellulosic biomass, macro algae lack lignin but contain a much broader range of polysaccharides. Enzymes for the degradation of polysaccharides found in macro algae such as agar, alginate, fucoidan, laminarin, and ulvan are not available. Solutions for the degradation of xylan and cellulose, however, are commercially available. The aim of the work has been to bioprospect geothermally-heated intertidal pools north of Húsavík for microbes producing 1,2-propanediol and/or glycohydrolases capable of degrading the polysaccharides found in macro algae. The temperature in the sampled geothermal pools ranged from 22 to 67°C. Samples were collected and enriched on various polysaccharides (laminarin, ulvan, fucoidan, agar, alginate, cellulose) as well as single carbohydrates (glucose, L-rhamnose, and L-fucose). A total of 195 isolates were obtained and characterized using standard phenotypic methods including API 20E, API ZYM panels and 16S rRNA. 1,2-PD was produced by 68 of the isolates from either glucose, rhamnose, or fucose. Enzymatic activities of the isolates are yet to be confirmed. 1 Introduction 3 Results and Discussion 3.1 Isolation and Characterization Macro algae species are potential feed-stocks for biofuel and fine chemical production Contain diverse polysaccharides Lack of commercially available enzymes for degradation is a major barrier Aim of this work is to Isolate glycohydrolases for macro algae processing Produce 1,2-propanediol with bacteria 195 isolates Majority of 1,2-PD producing isolates were enteric pathogens 1 isolate from 65° (B. licheniformis) 68 1,2-PD producers 117 VP positive strains Isolated strains were characterized by sequencing of the 16S rDNA gene. Only 1,2-PD producers were sequenced 2 Materials and Methods Samples were collected from six geothermal intertidal pools 3.2 Glycohydrolases and Other Enzymes 46+ strains degrade starch 26+ strains degrade β-glucan 51+ strains mobilize inorganic phosphate Work on algal polysaccharide degradation is ongoing (agar, alginate, fucoidan, ulvan, laminarin, etc) The pools north of Húsavík ranged in temperature from 23 to 65°C. Macro algae debris that has washed into the pool is visible 3.3 Evaluation of 1,2-Propanediol Production 20 of the 1,2-PD positive strains were screened at various concentrations of glucose & rhamnose (10 to 60 mM) Selected strains were grown on glu, rha, glu+rha, and 1,2-PD kinetically Palmaria palmata – a source of alginate and agar Ulva intestinalis – a source of ulvan Enrichment Medium Target Enzyme or Product Cellulose Cellulase Fucoidan Fucosidase Ulvan Ulvanase Laminarin Laminarase Alginate Alginate lyase Agar Agarase Tryptone Proteases Chitin Chitinases L-Fucose 1,2-Propanediol L-Rhamnose D-Glucose Non-specific Samples enriched 3x Time (h) Time (h) Shigella sonnei strain D1A grown on L-rhamnose (20 mM, left) and glu+rha (20 mM each, right) 3.4 Future Directions Isolates characterized by 16S rDNA and screened for 1,2-PD production and glycohydrolase production Further characterization by API 20E, API ZYM, and API 50CH Bioprocessing of macro algae Fermentation of macro algae hydrolysates for the production of high-value products