Fig. 7.
The possibilities of substrate channeling and superchanneling by targeting complexes of the Arg/N-degron pathway. Shown here are diagrams for reactions mediated by the S. cerevisiae Arg/N-degron–targeting complex (Fig. 6B). The setting of the human complex (Fig. 6A) is similar. To simplify diagrams, we left aside (as discussed in Results and Discussion) the scUFD4–scUBC4/5 part of the yeast targeting complex (scUFD4 E3 is not an N-recognin). (A) Substrate channeling. Nt-residues are denoted by single-letter abbreviations. Yellow ovals denote the rest of a protein substrate. A chain of black ovals is a substrate-linked poly-Ub chain. The hypothetical possibility of substrate channeling is diagrammed as a sequence of reaction/relocation steps for an (initially) Nt-Asn-bearing substrate. The substrate is sequentially Nt-deamidated and Nt-arginylated, then relocates and binds, via its conjugated Nt-Arg residue, to scUBR1–scRAD6 E3-E2, and is polyubiquitylated by this Ub ligase, followed by the proteasome-mediated degradation of substrate. The diagram uses notations of transition probabilities, at each reaction/relocation step, for a molecule of an (initially) Nt-Asn-bearing substrate. P1r, P2r, and P3r denote the probabilities of substrate moving through each reaction/relocation step while being retained (“r”) within the targeting complex (Fig. 6B). P1d, P2d, and P3d denote the “alternative” probabilities of substrate’s dissociation (”d”), at each reaction/relocation step, into the bulk solution. An efficacious (as distinguished from weak or nonexistent) substrate channeling would obtain, in this setting, if P1d << P1r, P2d << P2r, and P3d << P3r. (B) Superchanneling. In the hypothetical concept of superchanneling (see Results and Discussion), the transitions (and associated probabilities) that are shown in A are “supplemented” with probabilities of superchanneling (“sc”) transitions (P1sc, P2sc, and P3sc) at each reaction/relocation step. These transitions would result, at each step, in a bypass of downstream steps and direct polyubiquitylation of the substrate. In the language of probability-based transitions, an efficacious superchanneling would obtain, for example, if P1r << P1sc and P1d << P1sc; that is, if the initial binding of an Nt-Asn–bearing substrate to the complex-bound scNTA1 Nt-amidase would be rapidly followed by substrate’s polyubiquitylation, without the necessity of (mechanistically independent) Nt modifications and accompanying physical relocations of the substrate. Superchanneling would not occur if P1sc, P2sc, and P3sc are negligible in comparison to corresponding P1r and P1d probabilities.