BioPHYSICAL chemistry

CHAPTER 19 MOLECULAR IMAGING 409

A

B

0.2

0.4

0.6

0.8

1.0

(a)

0.0

300 350 400 450 500 550

Absorbance

Wavelength (nm)

0.2

0.4

0.6

0.8

1.0

(b)

0.0

56789

Normalized fluorescence

pH

Figure 19.5The pH dependence of the optical spectrum of YFP. (a) The amplitudes of the
two absorption bands have a distinct pH dependence with an isosbestic point. (b) The relative
amplitude can be fit to a pH dependence with a pKAof 7. Modified from Elsliger et al. (1999).

O

O

cyclization

 H 2 O

 H 2 O

dehydrogenation

second

dehydrogenation

DsRed

chromophore

GFP

chromophore

HO

R

O

NH

O (^2) O
O
HO
N
N
R
O
NH
O
O
HO
N
N
R
O
N
O
O
HO
N
N
R
O 2
O
NH
N
NH
H
O
Figure 19.6A general scheme for chromophore formation in GFPs consisting of a cyclization step
followed by dehydration. In DsRed, the chromophore undergoes an additional step that results in a
longer conjugated system. Modified from Matz et al. (2003).