The gain of rod phototransduction: reconciliation of biochemical and electrophysiological measurements.
We have resolved a central and long-standing paradox in understanding the amplification of rod phototransduction by making direct measurements of the gains of the underlying enzymatic amplifiers. We find that under optimized conditions a single photoisomerized rhodopsin activates transducin molecules and phosphodiesterase (PDE) catalytic subunits at rates of 120-150/s, much lower than indirect estimates from light-scattering experiments. Further, we measure the Michaelis constant, Km, of the rod PDE activated by transducin to be 10 microM, at least 10-fold lower than published estimates. Thus, the gain of cGMP hydrolysis (determined by kcat/Km) is at least 10-fold higher than reported in the literature. Accordingly, our results now provide a quantitative account of the overall gain of the rod cascade in terms of directly measured factors.
Leskov
I B
IB
Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts 02114, USA.
Klenchin
V A
VA
Handy
J W
JW
Whitlock
G G
GG
Govardovskii
V I
VI
Bownds
M D
MD
Lamb
T D
TD
Pugh
E N
EN
Arshavsky
V Y
VY
D000818:Animals; D001001:Anura; D020134:Catalytic Domain; D006152:Cyclic GMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004594:Electrophysiology; D004789:Enzyme Activation / Q000187:drug effects; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D008027:Light; D008954:Models, Biological; D010727:Phosphoric Diester Hydrolases / Q000737:chemistry / Q000378:metabolism; D017948:Retinal Rod Photoreceptor Cells / Q000378:metabolism*; D012243:Rhodopsin / Q000737:chemistry / Q000378:metabolism; D012374:Rod Cell Outer Segment / Q000378:metabolism / Q000648:ultrastructure; D015293:Transducin / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D014785:Vision, Ocular / Q000502:physiology* / Q000528:radiation effects
D016244:Guanosine 5'-O-(3-Thiotriphosphate); D012243:Rhodopsin; D010727:Phosphoric Diester Hydrolases; D015293:Transducin; D006152:Cyclic GMP