Interaction of Cathelicidin LL-37 with human surfactant protein D (SP-D)
Contact: Dr. Edwin Veldhuizen. 030-535361, E.J.A.Veldhuizen@uu.nl.
Dept. Infectious Diseases & Immunology. Faculty of Veterinary Medicine.
Background
Vertebrates are constantly threatened by the invasion of microorganisms and have evolved
systems of immune defense to eliminate infective pathogens in the body. For the naïve host,
the primary antimicrobial defenses in the lung are the resident alveolar macrophages and
protein components including the cell wall-degrading enzyme lysozyme, the lung surfactant
and the small antimicrobial peptides. In order to study the first event that occur when
microbes land on the lower respiratory tract of the host, we need to study the interaction of
these innate immune proteins. In this project, the physical interaction between the human
surfactant protein D (hSP-D) and the human LL-37 will be performed using various kinds of
techniques such as Co-Immunoprecipitation, Size exclusion chromatography and ELISA. The
synergistic effect of the two innate immune proteins will also be studied using antimicrobial
activity assays (e.g. colony counting assay). Furthermore, other kind of small microbial
peptides can be included in the project.
The project
The student will work on his/her own project within an enthusiastic group of people with
similar research interests. The human SP-D will be purified from recombinant HEK293 cells
using Mannan affinity chromatography and size exclusion chromatography equipped with
Akta purifier. The interactions between LL-37 and human SP-D will be tested on various
methods such co-Immunoprecipitation, ELISA and also size exclusion chromatography. The
synergistic effect will be determined by antimicrobial activity tests using both gram negative
and positive bacteria.
Methods
Protein biochemistry:
Purification
Size exclusion chromatography (Akta)
SDS/PAGE analysis: western blot and Gel staining
Co-Immunoprecipitation
ELISA
Microbiology:
Antimicrobial activity tests: colony counting and bioscreen assay.