TY - JOUR
T1 - A soluble form of bovine rod photoreceptor phosphodiesterase has a novel 15-kDa subunit
AU - Gillespie, P. G.
AU - Prusti, R. K.
AU - Apel, E. D.
AU - Beavo, J. A.
N1 - Copyright:
Copyright 2004 Elsevier B.V., All rights reserved.
PY - 1989
Y1 - 1989
N2 - A substantial fraction (20-30%) of the bovine rod outer segment phosphodiesterase (PDE) activity is not associated with outer segment membranes prepared with buffers of moderate ionic strength; this PDE activity appears to represent a distinct, soluble isozyme. Although this PDE isozyme can be demonstrated to be present in sealed rod outer segments, it is discarded from most standard rod outer segment preparations. A method was developed that allowed the rapid purification of the soluble rod PDE by 2600-fold, to apparent homogeneity, using a monoclonal antibody column (ROS-1a). The soluble rod PDE isozyme has a novel M(r) = 15,000 subunit (δ) in addition to subunits of M(r) = 88,000 (α(sol)), 84,000 (β(sol)), and 11,000 (γ(sol)). The δ subunit comigrates with and may be identical to the cone PDE 15-kDa subunit. The small subunits of the soluble rod PDE and the membrane-associated rod PDE were isolated by reverse-phase chromatography. The γ(sol) subunit was a potent inhibitor of trypsin-activated rod PDE, inhibiting 50% of 1 pM PDE activity at a concentration of 11 pM. This concentration was similar to that observed for the γ subunit of the membrane-associated rod PDE. The purified δ subunit did not appear to affect PDE activity; this subunit was, however, unusually difficult to keep in solution. All of the kinetic and physical properties of the soluble rod PDE tested thus far are similar to those of the membrane-associated form, except for the presence of the δ subunit, suggesting that this unique subunit could mediate the solubility of the soluble rod PDE and the cone PDE in the intact photoreceptor.
AB - A substantial fraction (20-30%) of the bovine rod outer segment phosphodiesterase (PDE) activity is not associated with outer segment membranes prepared with buffers of moderate ionic strength; this PDE activity appears to represent a distinct, soluble isozyme. Although this PDE isozyme can be demonstrated to be present in sealed rod outer segments, it is discarded from most standard rod outer segment preparations. A method was developed that allowed the rapid purification of the soluble rod PDE by 2600-fold, to apparent homogeneity, using a monoclonal antibody column (ROS-1a). The soluble rod PDE isozyme has a novel M(r) = 15,000 subunit (δ) in addition to subunits of M(r) = 88,000 (α(sol)), 84,000 (β(sol)), and 11,000 (γ(sol)). The δ subunit comigrates with and may be identical to the cone PDE 15-kDa subunit. The small subunits of the soluble rod PDE and the membrane-associated rod PDE were isolated by reverse-phase chromatography. The γ(sol) subunit was a potent inhibitor of trypsin-activated rod PDE, inhibiting 50% of 1 pM PDE activity at a concentration of 11 pM. This concentration was similar to that observed for the γ subunit of the membrane-associated rod PDE. The purified δ subunit did not appear to affect PDE activity; this subunit was, however, unusually difficult to keep in solution. All of the kinetic and physical properties of the soluble rod PDE tested thus far are similar to those of the membrane-associated form, except for the presence of the δ subunit, suggesting that this unique subunit could mediate the solubility of the soluble rod PDE and the cone PDE in the intact photoreceptor.
UR - http://www.scopus.com/inward/record.url?scp=0024397052&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=0024397052&partnerID=8YFLogxK
M3 - Article
C2 - 2545702
AN - SCOPUS:0024397052
SN - 0021-9258
VL - 264
SP - 12187
EP - 12193
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 21
ER -