Current Projects and People involved
Summary Profile of Projects 2005/6
Members of the Centre were involved in a total of 9 research projects which were complete prior to 2005. The majority of current projects involve collaboration with DCU and NUI Maynooth through membership of the National Institute for Cellular Biotechnology and Tallaght Hospital (AMNCH).
Lead Researchers: M. Costello & J. Behan
Post Doctoral Fellow: Dr. Laura Oakey
This project centered on the antimicrobial peptide produced by Bifidobacterium longum ITT13. This follow-on project centered on the optimization of the purification regime used to purify the peptide in order to maximize its recovery. The bacterium Bifidobacterium longum ITT13 was shown to reproducibly produce antimicrobial activity. This 3 year project was funded by the HEA PRTLI Cycle 3 funds secured with DCU to set up the NICB and wound up in late 2005.Purification is being optimised to improve recovery as part of a follow-on project.Publications are in preparation.
Lead Researchers: M. Costello & J. Behan
PhD student: Pamela Morris
This project followed on from an earlier PhD project supervised by the lead researchers which resulted in the isolation of a new peptide produced by Staphylococcus warneri ITT 35. It involves research on the characterization and purification regime for the peptide. Detailed characterization studies of the peptide have already been completed and significantly the purified peptide has antimicrobial activity against one of the key cystic fibrosis pathogens. This discovery was made possible through internal collaboration with the cystic fibrosis research team in the college. Molecular studies including polymerase chain reactions (PCR) have proven that S. warneri ITT 35 and the cured strain of S. warneri ITT 35 does not contain the nukA and nukM genes. These two genes are indicative of genes found in the nukacin ISK-1 producing strain S. warneri JCM11004. This result would discount that ITT 35 is producing the same antimicrobial peptide as the JCM11004 strain and in effect illustrate the antimicrobial peptide studied here is novel. The project was funded by PRDSP Strand 3 funding awarded in 2001 and is close to completion.
Lead Researcher: S. McClean
Sam Maher PhD student:
This ongoing project involves the investigation of the ability of the antimicrobial peptide melittin to enhance paracellular permeability across the GI tract. This peptide has been examined with a view to exploiting it, or its analogues, in the delivery of poorly bioavailable drugs.Studies completed so far demonstrate that melittin has potential in the oral delivery of poorly absorbed drugs allowing less invasive administration techniques and aiding in increased patient compliance and cheaper delivery of biomolecules. A number of models have also been developed and evaluated to date:polarised Caco-2 cells, mucus secreting HT29 MTX E12 cells, a co-culture of both of these cell lines and an intact rat colonic epithelial model.Study of novel AMPs to examine any potential cytotoxicity in the existing eukaryotic cell systems have already developed. This project is funded by HEA Cycle 3 funds secured with DCU to set up the NICB.
Lead Researchers: J. Behan & M. Costello
MSc student: Margaret Morris (recommended for graduation 2006)
This project focused on the purification of an antimicrobial peptide from Serratia marcescens. The bacterium is a Gram-negative rod and was verified to be Serratia marcescens using the Biolog identification system. Purification protocols were established for the peptide which involved culturing the bacteria in 1L fermentations. Subsequent separation by anion exchange chromatography resulted in the isolation of two components which have demonstrated antimicrobial activity against A.caviae, E.aerogenes and E.coli 10536. They have also shown activity against B.multivorans and B.cenocepacia two opportunistic pathogens in cystic fibrosis patients. This project is funded by HEA Cycle 3 funds secured with DCU to set up the NICB.
Lead Researchers: M.Costello & J. Behan
Research Assistant: Claire Burgess
This project studied the antimicrobial peptide produced by a Staphylococcus aureus ITT 330 isolate. The molecular weight of the large antimicrobial peptide/protein purified from a clinical isolate of S. aureus has been confirmed as being approximately 50 kDa by Mass Spec analysis. Purification protocols have been developed and biochemical characterisation of the peptide shows it retains activity over a widepH range. The purified bacteriocin shows antimicrobial activity against K. rhizophila, E. faecalis, B. cereus and E. coli. This project is funded by HEA Cycle 3 funds secured with DCU to set up the NICB.
Lead Researchers: J. Behan & M. Costello
Research Assistant: Cormac Walsh
This project, which is now complete, involved optimisation of the purification regime for the peptide produced by Staphylococcus isolate ITT 218. Antimicrobial activity has been demonstrated through fermentative growth of the microorganism in tryptone soya broth in shake-flash culture at 37 0C and 150 RPM agitation. The active substance has been purified using cationic exchange chromatography as the capture step, hydrophobic interaction chromatography as an intermediate step, and size exclusion chromatography as a polishing step. Activity has been demonstrated against hospital isolates of methicillin-resistant Staphylococcus aureus. This project which was funded through thePRDSP Strand 3 programme used the supporting resources of the National Pharmaceutical Education Centre. Since the project completed, the Research Assistant has been working on contract in the NPEC and has applied for admission to conduct a Masters part time here at the Institute under the supervision of the lead researchers where he will study the scale up of production and purification of this peptide.
Lead Researcher: F. Kelleher
MSc student: Keith O Prionsias
This project involved the design and synthesis of chemical derivatives of the antimicrobial peptide nisin. Nisin is an antimicrobial peptide produced by lactococcus lactis species, and has been used as a food preservative for over 40 years.1 It is very potent against a wide range of both gram-positive and gram-negative bacteria, having potency in the nanomolar range compared to the micromolar range for most other antimicrobial peptides. It has been shown to have potent antibacterial activity against MRSA.
Many studies have been undertaken on the mechanism of action of this unique anti-microbial peptide and a recent study has elucidated its mechanism. Currently we are synthesising analogues of lanthionine where the sulphur linker atom is being replaced by an oxygen, nitrogen or carbon atom. The incorporation of these derivatives into the structure of Nisin will lead to analogs with very different chemical and physical properties.
To date we have been successful in synthesising fully protected lanthionine and we are at an advanced stage in the synthesis of the nitrogen linked analog. Preparation of the oxygen linked analog has proved more difficult synthetically but again we are at an advanced stage in this synthesis. This project is funded by HEA Cycle 3 funds secured with DCU to set up the NICB.
Lead Researchers: M.Costello & J. Behan
MSc student: Laura Cleary
This PRDSP Strand 1 project commenced in January 2006 and has been investigating the production of an antimicrobial peptide by a Staphylococcus aureus isolate ITT 37. Initial work on the purification of the extracellular peptide using cation exchange resin are focussing on the trials at a number of different pH values in order to optimise the capture step. Initial studies using reverse phase chromatography have given very positive results in that the antimicrobial activity was eluted over three fractions only which is very encouraging.
