Table 1 FRET-FLIM measurements showing that PopP2 L369P/V370P, L371P/D372P, and K383A are affected in their ability to physically interact in planta with the C-terminal portion of RRS1-R.

From: Secondary-structure switch regulates the substrate binding of a YopJ family acetyltransferase

Donor

Acceptor

Ʈ (ns)a

Δt (ns)(b)

Sem(c)

N(d)

E (%)(e)

p value(f)

PopP2-CFP

—

2.8377

0.302

0.01290

60

  

PopP2-CFP

RRS1-RCterm-YFPv

2.5356

 

0.03059

61

10.65

3.42E−15

L369P-V370P-CFP

—

2.5750

−0.185

0.02366

40

  

L369P-V370P-CFP

RRS1-RCterm-YFPv

2.7600

 

0.02483

40

−7.18

7.23E−07

L371P-D372P-CFP

—

2.5353

−0.062

0.02911

59

  

L371P-D372P-CFP

RRS1-RCterm-YFPv

2.5968

 

0.03179

60

−2.43

0.15615099

R380A-CFP

—

2.7649

0.098

0.02189

39

  

R380A-CFP

RRS1-RCterm-YFPv

2.6672

 

0.02571

39

3.53

0.004988117

K383A-CFP

—

2.5619

−0.241

0.04220

40

  

K383A-CFP

RRS1-RCterm-YFPv

2.8028

 

0.02391

40

−9.40

3.94E−06

  1. aMean lifetime, Ʈ, in nanoseconds (ns). For each nucleus, average fluorescence decay profiles were plotted and fitted with exponential function using a non-linear square estimation procedure and the mean lifetime was calculated according to Ʈ = Σαiti2/Σαiti with I(t) = Σαie−t/ti, (b) Δt = ƮD − ƮDA (in ns), (c) standard error of the mean, (d) total number of measured nuclei, (e) % FRET efficiency: E = 1 − (ƮDA/ƮD), and (f) p value of the difference between the donor lifetimes in the absence and presence of acceptor (Student’s t test). The statistical test used was two-sided. The lifetime measurements were carried out from two to three independent expression assays performed in N. benthamiana (leaf samples were taken between 36 and 48 h after infiltration with A. tumefaciens).
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