Figure 2
From: Biological insertion of computationally designed short transmembrane segments
Integration of computationally designed TM segments into microsomal membranes.
(A) Schematic of the engineered leader peptidase (Lep) model protein. Lep, consisting of 2 TM segments (H1 and H2) and a large luminal domain (P2), inserts into rough microsomes in an Nlum-C-lum orientation. Computationally designed TM sequences were engineered into the P2 domain with flanking glycosylation sites (G1 and G2). For sequences that integrate into the membrane (green), only the G1 site is glycosylated (left), whereas both G1 and G2 are modified for sequences (red) that do not integrate into the membrane (right). (B) In vitro translation in the presence (+) or absence (−) of rough microsomes (RM) of computationally designed TM sequences of different length (l) composed of Leu and Ala residues (top) and of Leu, Ala, Val, Ile, Gly, Phe, Thr, Ser, Met, Tyr, Trp, Pro, Asn and Arg residues with position-defined constraints (bottom). Non-glycosylated protein bands are indicated by an empty dot; singly and doubly glycosylated proteins are indicated by one or two black dots, respectively.
