Opdateres

Pig intestine mucosa contains unfractionated heparin, which can be extracted, refined and utilized for blood anticoagulant medicine. It is a widely used medical treatment for patients with a range of different medical conditions where the risk of blood clotting needs to be diminished. The heparin constituents are extracted from pig intestine mucosa collected at slaughterhouses in Denmark or abroad. A preservative agent is added to the mucosa material before it is shipped to a LEO Pharma production plant where the heparin is extracted and refined to the pharmaceutical product. However, during transportation and storing, a considerable proportion of the heparin is disappearing due to unknown physicochemical or biological mechanisms. The latter would probably imply enzymatic degradation involving enzymes from the mucosa tissue or from live or dead microbes. Although, the mucosa material is added antimicrobial chemicals, we know that some (unknown) bacteria can survive and potentially contribute to the degradation of the heparin. Any loss of heparin constitutes a challenge in terms of economical loss as well as spill of resources. Decreasing the loss of heparin in the feedstock mucosa will support LEO Pharma’s effort to make the entire process as sustainable as possible from a circular bioeconomic viewpoint.

Main objectives:

The project aims at clarifying the degradation mechanisms responsible for loss of heparin in the mucosa feedstock material. Specific research questions:

1. is the degradation of the heparin, in the mucosa, dependent on live microorganisms derived from the gut content?
2. which microorganisms are present in the mucosa at different stages of storage/transport?
3. can we suggest approaches to diminish the heparin degradation rate – e.g. enhanced inhibition of microbial proliferation or specific enzymatic processes?

Methodologies:

The master student will apply are range of approaches to clarify the research questions. Besides traditional microbial lab techniques, 16S DNA amplicon sequencing will be used to characterize the microbial community in the mucosa. Different biocidal agents will be employed to stop microbial proliferation/activity eventually responsible for heparin degradation. Advanced chemical analytical approaches, present at the department, may be utilized to characterize degradation metabolites and pathways.

Project setup:

The project is carried out in the microbiology section at Department of Environmental Science, Aarhus University, in close collaboration with LEO Pharma A/S company. The student can start at autumn semester 2024, but the starting time can be negotiated. Supervisors are senior scientists Anders Johansen and Niels Bohse Hendriksen, Section of Microbial Ecology, campus Roskilde.  

Contact: ajo@envs.au.dk; nbh@envs.au.dk

Department of Environmental Sciences, Frederiksborgvej 399, 4000 Roskilde

Microbial Biotechnology:

Cold-active enzymes from Arctic microorganisms for the Green Transition.

In many industrial processes, enzymes are attractive, green alternatives to chemicals. Today, enzymes are key ingredients in many products like food, textiles, pharmaceuticals, and detergents, and the enzyme market for detergents alone has become a multibillion dollar industry worldwide.

A major goal of the green transition strategy is CO2 reduction, which requires energy savings throughout society. A significant contribution to this goal will be a reduction of washing temperatures from current levels of 30-60°C down to 5-20°C. This can save up to 25.5 billion kWh/year in Europe, which in terms of CO2 emission equals approx. 18 million tons of CO2 yearly, corresponding to the annual emissions of 4 million cars.

For decades, it has been a goal for producers of detergent enzymes to develop cold-active enzymes due to their performance at low-temperature resulting in significant reduction in energy consumption and costs. However, the so-called cold-active detergent enzymes on the market today cannot be classified as truly cold-active, as their optimal activity is at temperatures above 30°C.

So, in order to achieve this reduction, truly cold-active enzymes for detergents are urgently needed to make washing efficient at low temperatures. The use of cold, but effective washes in households across Europe will be symbolic of the transition from more wasteful technologies and practices to greener and more sustainable ways of living.

Development of cold-active enzymes is currently approached in two ways: 1) Protein engineering of well-known commercial mesophilic enzymes or 2) to search for new truly cold-active enzymes in microorganisms from permanently cold environments.

The focus of this project is discovery of novel, truly cold-active enzymes. Besides being cold-active, enzymes for detergents also need to be active at high pH due to detergent pH (most often ≥ pH 9). The combination of low temperature and high pH is rare among natural environments, which limits the search for novel enzymes with both traits. However, one such environment exists, namely the ikaite columns in the Ikka Fjord in South Greenland.

The project will be a part of an ongoing project, ColdZymes, funded by DFF. Depending on the duration of the student project and personal interests, the student will be able to take part in one or more of the projects or tasks described below. 

The aim of either project is to identify and characterize detergent-relevant enzymes, which are active at low temperatures and high pH.

Project 1: Function-based screening for novel enzymes

In project 1, several microbiological and biotechnological techniques will be used.

Methods:

• Culturing of microorganisms on substrates mimicking the ikaite-environment
• Screening for novel enzymes using chromogenic substrates
• Characterization of microorganisms and enzymes
• Genomic analyses of cultured bacteria for identification of enzyme encoding genes
• Cloning and characterization of enzyme encoding genes (sequencing, genomics, bioinformatics).
• Heterologous expression, purification and characterization of enzymes

Project 2: Sequence-based screening for novel enzymes

Project 2 is mainly a bioinformatics project, but biotechnological techniques can be brought into play.

Methods:

• Isolation of environmental DNA from the ikaite columns
• Preparation of libraries for sequencing
• Genomic and metagenomic sequence analysis
• Cloning and characterization of enzyme encoding genes
• Heterologous expression, purification and characterization of enzymes

 Supervisors: Mariane Schmidt Thøgersen (Scientist, microbiology and biotechnology), mst@envs.au.dk and Thanassis Zervas (PostDoc, bioinformatics), az@envs.au.dk  

- April 2022

Microbial Biotechnology:

Enzymes degrading algal polysaccharide for pharmaceutical applications.

Marine polysaccharides like agar, carrageenan, fucoidan, and ulvan are extracted from a range of different species of macroalgae. As opposed to polysaccharides from land plants (e.g. cellulose, xylan, and pectin), the marine polysaccharides are sulfated, i.e. several saccharides in the chain carries a sulfate-group.

These polysaccharides have unique functional features like gelling, thickening, stabilizing, binding, and emulsifying effects of food.