Figure 5.

Trypsin Interacts with a Di-acidic Motif on the Extracellular Domain of PepT1 and PepT2

(A) SPR analysis of the MmPepT1^ECD interaction with trypsin. Inset: SPR sensorgram used to determine the binding constant. RU, response units. Error bars show the SEM (n = 3).

(B) The binding experiment in (A) was repeated with the RnPepT2^ECD protein.

(C) MST binding analysis reveals no interaction with the Gly-Sar peptide and abolition of trypsin interaction in the presence of high salt.

(D and E) Surface representation of (D) MmPepT1^ECD and (E) RnPepT2^ECD with the sequence conservation from cow, dog, chicken, human, mouse, and rat species mapped from blue to red. A highly conserved patch (indicated by the white dashed ellipse) was identified. Insets: MST binding analysis reveals an important role for D550 and E573 in MmPepT1^ECD, and D576 and E599 in RnPepT2^ECD, in mediating the electrostatic interaction with trypsin.