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. 2021 May 26;49(11):6489–6510. doi: 10.1093/nar/gkab414

Figure 5.

Figure 5.

Structure-based mutational analysis of the CCR4-CAF1 complex. (A) Effects of peripheral (non-active site) mutations on deadenylation by CCR4-CAF1 under standard reaction conditions (pH = 7.5). Replacement of positively charged surface residues on the CCR4 LRR domain with residues corresponding to the Sc CCR4 LRR domain delays deadenylation (R65E/+ or R87T/+). Deletion of the CCR4 N-terminal extension does not have a detectable effect (ΔN/+), whereas deletion of the CAF1 C-terminal tail facilitates deadenylation (+/ΔC). (B) Effects of peripheral (non-active site) mutations on deadenylation by CCR4-CAF1 under moderately acidic reaction conditions (pH = 6.0). Results are comparable to (A) without differential effects on either one of the nucleases. (C) Differential effect of a CCR4 active site mutation on deadenylation by CAF1. In contrast to a D491A mutation of the CCR4 active site (Figure 4D, –/+), a D491N mutation causes the CCR4-CAF1 complex to lose activity at moderately acidic pH and to display a pH dependence comparable to the isolated CAF1 nuclease (Figure 4F).