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. Author manuscript; available in PMC: 2022 Apr 5.
Published in final edited form as: Cell Rep. 2022 Mar 1;38(9):110436. doi: 10.1016/j.celrep.2022.110436

Figure 2. Sequence-guided mutations at the V2 hotspot region improve properties of well-formed trimmers.

Figure 2.

(A) V2 hotspot (V2-HS) region (res. 166–173) of clade C consensus sequence (n = 22,415) with corresponding C.1086 region mentioned below. C.1086 residues differing from the consensus sequence are in red.

(B) Influence of indicated V2-HS modifications in C.1086 UFO-v2 protein on binding to various mAbs, monitored by ELISA. The data are the average of more than two independent experiments.

(C) Trimeric proportion of C.1086 UFO-v2 (left) and BG505 SOSIP.664 V2-HS (right) variants; mean (value indicated) ± SD (error bars) estimated from SEC profiles of at least two independent purifications. *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001 (Student’s t test, two-tailed), (left) p value indicated on top of bar denotes comparison with UFO-v2, which has 166K,170H,173H.

(D) Blue native PAGE (BN-PAGE) of purified C.1086 proteins with molecular weight standard.

(E) BLI responses of different C.1086-purified designs (400 nM each) against V1/V2 trimer apex-specific PGT145 bnAbs.

(F) Comparison between binding affinities of optimized UFO-v2 V2-HS mutants relative to UFO-v2 against various env-specific mAbs. *KonUFO-v2/Kon UFO-v2 V2-HS for PGT121, as KD could not be calculated due to no observable dissociation. A fold change in affinity within 3-fold range is shaded gray and not considered a significant change.