Regulation of the HTRA2 protease activity by PDZ domain-binding peptide ligands and the influence on HTRA2-specific protein interaction networks in retinal tissues

ABSTRACT: For the identification of HTRA2-specific protein interaction partners in retinal tissues of the house swine (Sus scrofa), we performed co-immunoprecipitation experiments with recombinant 6His-tag HTRA2 followed by quantitative LC-MS-based proteomics (see Fig. 2A and File S1). In brief, 2 µg recombinant 6His-tag HTRA2 (HTRA2 group) were spiked into 5 mg homogenized porcine retina and was subsequently enriched with the bounded protein interaction partners via Ni-NTA affinity purification (n=3). In addition, two further sample groups were treated with 10 µg UCF 101 (UCF-101 group) or 10 µg synthetic CDR peptide (CDR group) to unravel the influence of both molecules on the HTRA2-specific protein interactome in the retinal tissues (n=3 for each group). Eluate fractions of each group were separated by 1-D SDS PAGE (Fig. S1) and subsequently subjected to in-gel trypsin digestion. To distinguish unspecific from HTRA2 specific binders, we also included a control Ni-NTA bead group (CTRL group) for the quantitative MS analysis (n=3).

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human) Sus Scrofa Domesticus (domestic Pig)

TISSUE(S): Retina

DISEASE(S): Open-angle Glaucoma

SUBMITTER: Carsten Schmelter  

LAB HEAD: Dr. Carsten Schmelter

PROVIDER: PXD024442 | Pride | 2021-10-11

REPOSITORIES: Pride