Characterization of PlyAZ3aT
Typ
Examensarbete för masterexamen
Master Thesis
Master Thesis
Program
Biotechnology (MPBIO), MSc
Publicerad
2018
Författare
Sundström, Clara
Modellbyggare
Tidskriftstitel
ISSN
Volymtitel
Utgivare
Sammanfattning
Antibiotic resistance among bacteria is a growing problem all around the world. Hence, new alternatives to antibiotics are demanded for. Bacteriophages (phages) represent such an alternative, as they are viruses that naturally affect bacteria. After infecting a bacterium, the phage uses enzymes to escape the host cell and kill it. Endolysins are one group of enzymes used in this step to cut open the bacterial peptidoglycan. A future perspective on antimicrobial agents is the direct usage of endolysins to treat bacterial infections. In the lab of Grégory Resch (Lausanne University, Switzerland) a new endolysin, PlyAZ3aT, was identified within the genome of Streptococcus tigurinus AZ3aT and its activity was evaluated in a series of experiments. In this report the explorative work on PlyAZ3aT have been continued by means of comparison with the formerly known endolysin Cpl-1 and chimeric enzymes (chimeras) constructed by domain swapping between the two native enzymes (PlyAZ3aT and Cpl-1). Six chimeras were purified from crude extract using Fast Protein Liquid Chromatography (FPLC). Thereafter, the activity of all enzymes was evaluated in vitro on S. pneumoniae D39 and S. tigurinus AZ3aT using turbidity, Minimal Inhibitory Concentration (MIC) and time-kill experiments. While all enzymes showed similar activity in poor medium (NaCl 0.9%), enzymes harbouring the Cell Wall Binding Domain (CWBD) of PlyAZ3aT performed significantly better on both strains in rich medium. Whether these first interesting results reflect a negative medium effect on Cpl-1 or an activity improvement provided by the PlyAZ3aT CWBD on dividing cells remains to be determined. In addition, to further evaluate the potential of PlyAZ3aT and/or any of the chimeras as antimicrobial agent, their activity should be verified in vivo.
Beskrivning
Ämne/nyckelord
Mikrobiologi , Biokatalys och enzymteknik , Infektionsmedicin , Livsvetenskaper , Microbiology , Biocatalysis and Enzyme Technology , Infectious Medicine , Life Science