Figure 2
From: A proteolytic modification of AIM promotes its renal excretion
Assessment of the AIM cleavage site.
(a) LC-MS analysis of urine sAIM fragments after LysC or GluC digestion. Detected fragments were indicated by red lines (LysC digestion) and green lines (GluC digestion) in amino acid sequence of mAIM. The amino acids of the region where the cleavage site may be present are colored by blue. Three SRCR domains are indicated by boxes. (b) Culture supernatant of HEK293T cells expressing variant AIM (indicated by amino acid number) and urine collected from AIM−/− mice injected with rAIM (200 μg) (indicated by U) were immunoblotted with the Rab2 antibody in a non-reducing condition. The AIMLys264 and the AIMGly265 were present at a comparable position with the urine sAIM (indicated by allows). (c) The AIMLys264Ala and the AIM265GlyAla (200 μg each) were injected intravenously into AIM−/− mice and the urine collected after 1, 2, and 3 h was analyzed for AIM by immunoblotting. No sAIM was detected in urine from AIMLys264Ala injected mice, whereas it was detected in urine from AIM265GlyAla injected mice. rAIM (25 ng) and urine collected from AIM−/− mice injected with rAIM (200 μg) (indicated by U) are presented as controls.
