Перегляд вихідного коду сторінки Some Frightful Yet Still Revolutionary CASK Ideas

A 22 residue synthetic peptide within the E2 domain (FD1) (Figure?1A and Figure?2B), containing the putative active site of?APP, possessed [http://en.wikipedia.org/wiki/CASK CASK] ferroxidase activity that was ?40% that of APP695�� (Figure?2C). Mutational analysis of APP695�� (Figure?2A) and FD1 (Figures 2B�C2D) confirmed that disruption of the REXXE motif, by altering a single conserved amino acid (REWEN, ��E14N�� in Figures 2B and 2C), or substituting the homologous pentapeptide regions of APLP1 (REWAM, ��EE13/14AM�� in Figure?2D) or APLP2 (KEWEE, ��R10K�� in Figure?2D), abolished activity. Substitution with the homologous H-ferritin sequence (REHAE, ��WE12/13HA�� in Figure?2B), which does not disrupt the consensus motif, retained activity (Figure?2D). Like FD1 peptide (Figure?2C), purified E2 polypeptide [http://www.selleckchem.com/products/ulixertinib-bvd-523-vrt752271.html Ulixertinib purchase] (Figure?1A) possessed ?40% of the ferroxidase activity of APP695�� (Figure?2A). We explored for other domains of APP needed to restore full activity to E2. Whereas purified E1 domain possessed no ferroxidase activity, equimolar concentrations of E1 doubled E2 activity (Figure?2E) to about that of APP695�� (Figure?2A). We mapped this potentiation effect to the growth factor domain (GFD) within E1 (Rossjohn et?al., 1999) (Figure?2A). GFD did not engender activity from APLP2 (Figure?2A), consistent with the requirement for the REXXE motif. We hypothesized that APP ferroxidase activity may facilitate iron movement analogous to the interaction of CP with ferroportin (De?Domenico et?al., 2007). Ferroportin may be expressed in all cells, but CP is not, leading us to suspect that APP may play the ferroxidase role [http://www.selleckchem.com/JAK.html JAK inhibitor] in certain cells that lack CP such as HEK293T (De Domenico et?al., 2007) and cortical neurons (Klomp et?al., 1996). The impact of endogenous APP suppression by RNAi on iron export was therefore initially studied in HEK293T cells, where the absence of CP was confirmed by western blot (not shown). APP-suppressed cells accumulated significantly more (?50%, p