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Functions of the N-terminal region of cyclic nucleotide phosphodiesterase 3 (PDE 3) isoforms

Kenan, Y; Murata, T; Shakur, Y; Degerman, Eva LU and Manganiello, V C (2000) In Journal of Biological Chemistry 275(16). p.12331-12338
Abstract
The N-terminal portion of phosphodiesterase (PDE) 3 was arbitrarily divided into region 1 (amino acids 1-300), which contains a large hydrophobic domain with six predicted transmembrane helices, and region 2 (amino acids 301-500), with a smaller hydrophobic domain ( approximately 50 residues). To analyze these regions, full-length human (H)PDE3A and mouse (M)PDE3B and a series of N-terminal truncated mutants were synthesized in Sf9 cells. Activities of HPDE3A, H3A-Delta189, MPDE3B, and M3B-Delta196, which retained all or part of the hydrophobic domain in region 1, were recovered almost entirely in particulate fractions. H3A-Delta321 and M3B-Delta302, containing region 2, were recovered essentially equally in particulate and cytosolic... (More)
The N-terminal portion of phosphodiesterase (PDE) 3 was arbitrarily divided into region 1 (amino acids 1-300), which contains a large hydrophobic domain with six predicted transmembrane helices, and region 2 (amino acids 301-500), with a smaller hydrophobic domain ( approximately 50 residues). To analyze these regions, full-length human (H)PDE3A and mouse (M)PDE3B and a series of N-terminal truncated mutants were synthesized in Sf9 cells. Activities of HPDE3A, H3A-Delta189, MPDE3B, and M3B-Delta196, which retained all or part of the hydrophobic domain in region 1, were recovered almost entirely in particulate fractions. H3A-Delta321 and M3B-Delta302, containing region 2, were recovered essentially equally in particulate and cytosolic fractions. H3A-Delta397 and H3A-Delta457, lacking both hydrophobic domains, were predominantly cytosolic. H3A-Delta510 and M3B-Delta604, lacking both regions 1 and 2, were virtually completely cytosolic. M3B-Delta196 eluted as a large aggregated complex during gel filtration. With removal of greater amounts of N-terminal sequence, aggregation of PDE3 decreased, and H3A-Delta607, H3A-Delta721, and M3B-Delta604 eluted as dimers. Truncated HPDE3A proteins were more sensitive than full-length HPDE3A to inhibition by lixazinone. These results suggest that the hydrophobic domains in regions 1 and 2 contain structural determinants important for association of PDE3 with intracellular membranes, as well for self-association or aggregation during gel filtration and sensitivity to a specific inhibitor. (Less)
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author
organization
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type
Contribution to journal
publication status
published
subject
in
Journal of Biological Chemistry
volume
275
issue
16
pages
12331 - 12338
publisher
ASBMB
external identifiers
  • pmid:10766874
  • scopus:0034697174
ISSN
1083-351X
language
English
LU publication?
yes
id
7d44a575-5710-4f07-a45f-31fd991c105c (old id 1117959)
alternative location
http://www.jbc.org/cgi/content/abstract/275/16/12331
date added to LUP
2008-06-25 08:17:43
date last changed
2017-01-22 03:23:20
@article{7d44a575-5710-4f07-a45f-31fd991c105c,
  abstract     = {The N-terminal portion of phosphodiesterase (PDE) 3 was arbitrarily divided into region 1 (amino acids 1-300), which contains a large hydrophobic domain with six predicted transmembrane helices, and region 2 (amino acids 301-500), with a smaller hydrophobic domain ( approximately 50 residues). To analyze these regions, full-length human (H)PDE3A and mouse (M)PDE3B and a series of N-terminal truncated mutants were synthesized in Sf9 cells. Activities of HPDE3A, H3A-Delta189, MPDE3B, and M3B-Delta196, which retained all or part of the hydrophobic domain in region 1, were recovered almost entirely in particulate fractions. H3A-Delta321 and M3B-Delta302, containing region 2, were recovered essentially equally in particulate and cytosolic fractions. H3A-Delta397 and H3A-Delta457, lacking both hydrophobic domains, were predominantly cytosolic. H3A-Delta510 and M3B-Delta604, lacking both regions 1 and 2, were virtually completely cytosolic. M3B-Delta196 eluted as a large aggregated complex during gel filtration. With removal of greater amounts of N-terminal sequence, aggregation of PDE3 decreased, and H3A-Delta607, H3A-Delta721, and M3B-Delta604 eluted as dimers. Truncated HPDE3A proteins were more sensitive than full-length HPDE3A to inhibition by lixazinone. These results suggest that the hydrophobic domains in regions 1 and 2 contain structural determinants important for association of PDE3 with intracellular membranes, as well for self-association or aggregation during gel filtration and sensitivity to a specific inhibitor.},
  author       = {Kenan, Y and Murata, T and Shakur, Y and Degerman, Eva and Manganiello, V C},
  issn         = {1083-351X},
  language     = {eng},
  number       = {16},
  pages        = {12331--12338},
  publisher    = {ASBMB},
  series       = {Journal of Biological Chemistry},
  title        = {Functions of the N-terminal region of cyclic nucleotide phosphodiesterase 3 (PDE 3) isoforms},
  volume       = {275},
  year         = {2000},
}