Mole quantity of RPE65 and its productivity in the generation of 11-cis-retinal from retinyl esters in the living mouse eye

Arkady L. Lyubarsky, Andrey B. Savchenko, Sarah B. Morocco, Lauren L. Daniele, T. Michael Redmond, Edward N Pugh Jr

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

RPE65, a protein expressed in cells of the retinal pigment epithelium of the eye, is essential for the synthesis by isomerohydrolase of 11-cis-retinal, the chromophore of rod and cone opsins. Recent work has established that RPE65 is a retinyl ester binding protein, and as all-trans-retinyl esters are the substrate for isomerohydrolase activity, the hypothesis has emerged that RPE65 serves to deliver substrate to this enzyme or complex. We bred mice with five distinct combinations of the RPE65 Leu450/Met450 variants (Leu/Leu, Met/Met, Leu/Met, Leu/-, and Met/-), measured in mice of each genotype the mole quantity of RPE65 per eye, and measured the initial rate of rhodopsin regeneration after a nearly complete bleach of rhodopsin to estimate the maximum rate of 11-cis-retinal synthesis in vivo. The quantity of RPE65 per eye ranged from 5.7 pmol (Balb/c) to 0.32 pmol (C57BL/6N × Rpe65-/-); the initial rate of rhodopsin regeneration was a Michaelis function of RPE65, where V max = 18 pmol/min per eye and Km = 1.7 pmol, and not dependent on the Leu450/Met450 variant. At RPE65 levels well below the K m, the rate of production of 11-cis-retinal per RPE65 molecule was ∼ 10 min-1. Thus, the results imply that as a chaperone each RPE65 molecule can deliver retinyl ester to the isomerohydrolase at a rate of 10 molecules/min; should RPE65 itself be identified as the isomerase, each copy must be able to produce at least 10 molecules of ll-cis-retinal per minute.

Original languageEnglish (US)
Pages (from-to)9880-9888
Number of pages9
JournalBiochemistry
Volume44
Issue number29
DOIs
StatePublished - Jul 26 2005
Externally publishedYes

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Retinaldehyde
Rhodopsin
Esters
Productivity
Molecules
Regeneration
Pigment Epithelium of Eye
Rod Opsins
Cone Opsins
Retinal Cone Photoreceptor Cells
Retinal Rod Photoreceptor Cells
Isomerases
Vertebrate Photoreceptor Cells
Retinal Pigment Epithelium
Retinal Pigments
Carrier Proteins
Substrates
Chromophores
Genotype
Enzymes

ASJC Scopus subject areas

  • Biochemistry

Cite this

Mole quantity of RPE65 and its productivity in the generation of 11-cis-retinal from retinyl esters in the living mouse eye. / Lyubarsky, Arkady L.; Savchenko, Andrey B.; Morocco, Sarah B.; Daniele, Lauren L.; Redmond, T. Michael; Pugh Jr, Edward N.

In: Biochemistry, Vol. 44, No. 29, 26.07.2005, p. 9880-9888.

Research output: Contribution to journalArticle

Lyubarsky, Arkady L. ; Savchenko, Andrey B. ; Morocco, Sarah B. ; Daniele, Lauren L. ; Redmond, T. Michael ; Pugh Jr, Edward N. / Mole quantity of RPE65 and its productivity in the generation of 11-cis-retinal from retinyl esters in the living mouse eye. In: Biochemistry. 2005 ; Vol. 44, No. 29. pp. 9880-9888.
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AU - Daniele, Lauren L.

AU - Redmond, T. Michael

AU - Pugh Jr, Edward N

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