Expression and characterization of deletion recombinants of two cGMP-inhibited cyclic nucleotide phosphodiesterases (PDE-3)
(1998) In Cell Biochemistry and Biophysics 29(1-2). p.89-111- Abstract
- cDNAs encoding two PDE-3 or cyclic GMP-inhibited (cGI) cyclic nucleotide phosphodiesterase (PDE) isoforms, RPDE-3B (RcGIP1) and HPDE-3A (HcGIP2), were cloned from rat (R) adipose tissue and human (H) heart cDNA libraries. Deletion and N- and C-terminal truncation mutants were expressed in Escherichia coli in order to define their catalytic core. Active mutants of both RPDE-3B and HPDE-3A included the domain conserved among all PDEs plus additional upstream and downstream sequences. An RPDE-3B mutant consisting of the conserved domain alone and one from which the RPDE-3B 44-amino acid insertion was deleted exhibited little or no activity. All active recombinants exhibited a high affinity (< 1 microM) for cyclic AMP (cAMP) and cyclic GMP... (More)
- cDNAs encoding two PDE-3 or cyclic GMP-inhibited (cGI) cyclic nucleotide phosphodiesterase (PDE) isoforms, RPDE-3B (RcGIP1) and HPDE-3A (HcGIP2), were cloned from rat (R) adipose tissue and human (H) heart cDNA libraries. Deletion and N- and C-terminal truncation mutants were expressed in Escherichia coli in order to define their catalytic core. Active mutants of both RPDE-3B and HPDE-3A included the domain conserved among all PDEs plus additional upstream and downstream sequences. An RPDE-3B mutant consisting of the conserved domain alone and one from which the RPDE-3B 44-amino acid insertion was deleted exhibited little or no activity. All active recombinants exhibited a high affinity (< 1 microM) for cyclic AMP (cAMP) and cyclic GMP (cGMP), were inhibited by cAMP, cGMP, and cilostamide, but not by rolipram, and were photolabeled with [32P]-cGMP. The IC50 values for cGMP inhibition of cAMP hydrolysis were lower for HPDE-3A than for RPDE-3B recombinants. The deduced amino acid sequences of HPDE-3A and RPDE-3B catalytic domains are very similar except for the 44-amino acid insertion not found in other PDEs. It is possible that this insertion may not only distinguish PDE-3 catalytic domains from other PDEs and identify catalytic domains of PDE-3 subfamilies or conserved members of the PDE-3 gene family, but may also be involved in the regulation of sensitivity of PDE-3s to cGMP. (Less)
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https://lup.lub.lu.se/record/1113551
- author
- He, R ; Komas, N ; Ekholm, Dag ; Murata, T ; Taira, M ; Hockman, S ; Degerman, Eva LU and Manganiello, V C
- organization
- publishing date
- 1998
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Cell Biochemistry and Biophysics
- volume
- 29
- issue
- 1-2
- pages
- 89 - 111
- publisher
- Humana Press
- external identifiers
-
- pmid:9631240
- scopus:0031634909
- ISSN
- 1085-9195
- DOI
- 10.1007/BF02737830
- language
- English
- LU publication?
- yes
- id
- b6bdee3e-f905-449e-8b2b-010b111bbbee (old id 1113551)
- date added to LUP
- 2016-04-01 16:59:30
- date last changed
- 2022-01-28 23:33:57
@article{b6bdee3e-f905-449e-8b2b-010b111bbbee, abstract = {{cDNAs encoding two PDE-3 or cyclic GMP-inhibited (cGI) cyclic nucleotide phosphodiesterase (PDE) isoforms, RPDE-3B (RcGIP1) and HPDE-3A (HcGIP2), were cloned from rat (R) adipose tissue and human (H) heart cDNA libraries. Deletion and N- and C-terminal truncation mutants were expressed in Escherichia coli in order to define their catalytic core. Active mutants of both RPDE-3B and HPDE-3A included the domain conserved among all PDEs plus additional upstream and downstream sequences. An RPDE-3B mutant consisting of the conserved domain alone and one from which the RPDE-3B 44-amino acid insertion was deleted exhibited little or no activity. All active recombinants exhibited a high affinity (< 1 microM) for cyclic AMP (cAMP) and cyclic GMP (cGMP), were inhibited by cAMP, cGMP, and cilostamide, but not by rolipram, and were photolabeled with [32P]-cGMP. The IC50 values for cGMP inhibition of cAMP hydrolysis were lower for HPDE-3A than for RPDE-3B recombinants. The deduced amino acid sequences of HPDE-3A and RPDE-3B catalytic domains are very similar except for the 44-amino acid insertion not found in other PDEs. It is possible that this insertion may not only distinguish PDE-3 catalytic domains from other PDEs and identify catalytic domains of PDE-3 subfamilies or conserved members of the PDE-3 gene family, but may also be involved in the regulation of sensitivity of PDE-3s to cGMP.}}, author = {{He, R and Komas, N and Ekholm, Dag and Murata, T and Taira, M and Hockman, S and Degerman, Eva and Manganiello, V C}}, issn = {{1085-9195}}, language = {{eng}}, number = {{1-2}}, pages = {{89--111}}, publisher = {{Humana Press}}, series = {{Cell Biochemistry and Biophysics}}, title = {{Expression and characterization of deletion recombinants of two cGMP-inhibited cyclic nucleotide phosphodiesterases (PDE-3)}}, url = {{http://dx.doi.org/10.1007/BF02737830}}, doi = {{10.1007/BF02737830}}, volume = {{29}}, year = {{1998}}, }