Petrovicet al., Science 376 , eabm9798 (2022) 10 June 2022 3of18
Fig. 2. Structural and biochemical analyses of the Nup192-Nic96 and
Nup188-Nic96 interactions.(A) Domain structures ofC. thermophilumNup188,
Nup192, and Nic96. (B) Cartoon representation of the 3.8-ÅC. thermophilum
Nup192•Nic96R2single-particle cryo-EM structure. Inset regions are magnified to
illustrate the molecular details of the Nup192-Nic96 interaction. Red circles
indicate residues involved in the Nup192-Nic96 interaction. (C andD) Summary
of the effect of structure-guided (C) SUMO-Nic96R2and (D) Nup192 mutations
on Nup192•SUMO-Nic96R2complex formation, assayed by SEC. +++, no effect;
++, weak effect; +, moderate effect;–, abolished binding. (E) KDs determined by
triplicate ITC experiments, with the mean and associated standard error
reported. (F) SEC-MALS interaction analyses of Nup192•SUMO-Nic96R2and
interaction-abolishing mutants. Measured molecular masses are indicated,
with theoretical masses in parentheses. S, small ubiquitin-like modifier
(SUMO). (G) Two views in cartoon representation of the 4.4-ÅC. thermophilum
Nup188•Nic96R2crystal structure phased and built with the 2.8-Å Nup188NTD
and 3.4-Å Nup188Tail[Protein Data Bank (PDB) ID 5CWU] ( 28 ) crystal structures.
Inset regions are magnified to illustrate the molecular details of the Nup188-Nic96R2
interaction. Red circles indicate residues involved in Nup188-Nic96R2binding.
(H andI) Summary of the effect of structure-guided (H) SUMO-Nic96R2and (I)
Nup188 mutations on Nup188•SUMO-Nic96R2complex formation, assayed by
SEC. (J) KDs determined by triplicate ITC experiments with the mean and
associated standard error reported. (K) SEC-MALS analysis of Nup188•SUMO-
Nic96R2and interaction-abolishing mutants. Single-letter abbreviations for the
amino acid residues are as follows: A, Ala; C, Cys; D, Asp; E, Glu; F, Phe; G, Gly;
H, His; I, Ile; K, Lys; L, Leu; M, Met; N, Asn; P, Pro; Q, Gln; R, Arg; S, Ser; T,
Thr; V, Val; W, Trp; and Y, Tyr.RESEARCH | STRUCTURE OF THE NUCLEAR PORE