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Figure 3.
Figure 3. Enzyme regulation requires integrity of the linker
segment that forms the αN10-switch in the Rv1264 holoenzyme.
(a) Comparison of the region forming the αN10-switch in the
active state (PDB entry 1Y11, shown in green and ochre) and the
inhibited state (PDB entry 1Y10, shown in blue and red). The
overlay is based on superposition of the regulatory domains; the
catalytic domains are offset by 55° rotation and 6 Å
movement between active and inhibited conformations.^8 The
ribbon diagrams show with helix α10 starting at residue 160,
the linker with the αN10-switch, and the first β-strand (β1)
of the catalytic domains. The position of residue 204, where
additional amino acids were inserted in the mutagenesis study,
is indicated for active and inhibited states. Positions of
residues Met193 and Met194 that have been exchanged in the
earlier study are indicated.^8 (b) pH activity profiles of
insertion of one, two, three or nine amino acids at position 204
can be seen. Standard deviations are shown as vertical bars if
they exceed the symbol size, which corresponds to SD = 10%.
While one or two inserted amino acids ((●) +S204 and (○)
+SA204) result in an overall inhibited phenotype, insertion of
three residues ((▪) +SAA204) displays a phenotype similar to
the wild-type enzyme (broken line). Figure 3. Enzyme
regulation requires integrity of the linker segment that forms
the αN10-switch in the Rv1264 holoenzyme. (a) Comparison of the
region forming the αN10-switch in the active state (PDB entry
1Y11, shown in green and ochre) and the inhibited state (PDB
entry 1Y10, shown in blue and red). The overlay is based on
superposition of the regulatory domains; the catalytic domains
are offset by 55° rotation and 6 Å movement between
active and inhibited conformations.[3]^8 The ribbon diagrams
show with helix α10 starting at residue 160, the linker with
the αN10-switch, and the first β-strand (β1) of the catalytic
domains. The position of residue 204, where additional amino
acids were inserted in the mutagenesis study, is indicated for
active and inhibited states. Positions of residues Met193 and
Met194 that have been exchanged in the earlier study are
indicated.[4]^8 (b) pH activity profiles of insertion of one,
two, three or nine amino acids at position 204 can be seen.
Standard deviations are shown as vertical bars if they exceed
the symbol size, which corresponds to SD = 10%. While one or two
inserted amino acids ((●) +S204 and (○) +SA204) result in an
overall inhibited phenotype, insertion of three residues ((▪)
+SAA204) displays a phenotype similar to the wild-type enzyme
(broken line). Introduction of the nonapeptide SAAGPSGAA ((□)
+9aa-204) leads to a decoupling of regulatory and catalytic
domains, as this enzyme is, like the M193P/M194P mutant, no
longer pH-responsive.[5]^8
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