Timeclok.DBF: a microcomputer application.
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Microcomputers↗
Biomedical subjects
D R Patton
Publications and source records attributed to D R Patton.
Structure-activity relationships for the carboxy-terminus truncated analogues of angiotension II, a new class of angiotensin II antagonists.
A series of analogues of the recently reported angiotensin II (AII) antagonist [Sar1]AII-(1-7)-amide or des-Phe8[Sar1]AII (3) have been prepared by solid-phase synthesis and purified by reverse-phase liquid chromatography. The agonist and antagonist properties of these carboxy-truncated analogues of AII were determined in the isolated rabbit aorta assay. In the analogues tested, replacement of aspartic acid in position 1 by sarcosine was found necessary to produce significant antagonist activity. At position 7 of the des-Phe8 analogues, prolinamide could be replaced by proline without significant change in the biological activity. However, substitution of 7-prolinamide by either glycinamide or sarcosinamide provided inactive peptides. Methylation of the 4-tyrosine in [Sar1]AII-(1-7)-NH2 preserved the antagonist potency in isolated rabbit aorta. Deletion of the proline at position 7 resulted in inactive hexapeptides, both in the Asp1 and Sar1 series. However synthesis of the N,N-dimethyl amide at the N-terminus afforded hexapeptide [Sar1]AII-(1-6)-N(CH3)2 (10) with a pA2 value of 7.05. All the antagonistic peptides synthesized were fully reversible, competitive antagonists in vitro. These findings indicate that the structural requirements for receptor blockade by these C-truncated analogues are quite stringent with respect to the nature of the amino acid at positions 1 and 6/7. The analogues 2, 3, 7, 10, 11 (saralasin), and 12 (sarmesin) were tested in vivo in the anesthetized rat and were found to inhibit the AII pressor response. In addition, 3 inhibited angiotensin II stimulated aldosterone release from isolated rat adrenal zona glomerulosa cells and had no agonist activity by itself at the doses tested. Interestingly, analogue 3, when injected intracerebroventricularly in conscious rats, failed to antagonize the dipsogenic response to an angiotensin II icv injection and this reflects some heterogeneity in the AII receptor population. Peptide 3 is the first example of an antagonist that discriminates between peripheral and brain receptor subtypes.
Angiotensin II↗
Conformational restriction of angiotensin II: cyclic analogues having high potency.
Cyclic analogues of angiotensin II (AII) were synthesized by connecting the side chains of residues 3 and 5 via a disulfide bridge. Appropriate conformational constraints afforded an analogue, [Hcy3,5]AII, having high contractile activity (pD2 = 8.48 vs 8.81 for AII) and excellent binding affinity (IC50 = 2.1 nM vs 2.2 nM for AII). This type of cyclization was also used to prepare a highly potent AII antagonist, [Sar1,Hcy3,5,Ile8]AII (pA2 = 9.09 vs 9.17 for [Sar1, Ile8]AII; IC50 = 0.9 nM vs 1.9 nM for [Sar1,Ile8]AII). Model building suggests that this ring structure is consistent with a receptor-bound conformation having any of a variety of three-residue turns, including a gamma-turn. In contrast, the receptor-bound conformation of AII does not appear to accommodate a beta-turn or an alpha-helix which includes residues 3-5.
Amino Acid Sequence↗
Biophotography 1990--a survey of the field and its practitioners.
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Data Collection↗
A synthetic linear decapeptide binds to the atrial natriuretic peptide receptors and demonstrates cyclase activation and vasorelaxant activity.
A linear decapeptide, [cyclohexylalanine 106]ANP-(105-114)NH2 (1), where ANP is atrial natriuretic peptide, was prepared by solid phase synthesis and purified by reverse-phase liquid chromatography. This novel peptide was found to bind to ANP receptors in rabbit lung membranes, to stimulate cGMP production in various tissues, and to fully relax precontracted rabbit aorta in a dose-dependent fashion. The potency of 1 in the various in vitro assays varies between one-twentieth and one-eightieth of the potency of the reference peptide, the 24-mer rat ANP-(103-126). The linear decapeptide 1, which encompasses amino acid residues from the rat ANP sequence (105-114), features a cyclohexylalanine residue instead of the phenylalanine 106 residue in the hormone sequence, a free sulfhydryl function at the N-terminal cysteine 105, and a carboxamide C terminus. Its disulfide dimer 6 was active in the rabbit aorta assay while the S-methyl cysteine 7 analogue was not active in the same assay at similar concentrations. The decapeptide 1 is of particular significance because it is the shortest analogue reported to date endowed with agonistic activity at the guanylate cyclase-coupled ANP receptor. In particular, it is interesting to compare its structure to the structures of other short linear analogues of ANP which are totally devoid of the ability to stimulate particulate guanylate cyclase activity.
Adenylyl Cyclases↗
Role of anemia in the pathogenesis of left ventricular hypertrophy in end-stage renal disease.
Left ventricular (LV) hypertrophy is common in end-stage renal disease, yet the factors associated with its development are poorly understood. LV mass index was determined by echocardiography in 78 patients who had been treated by dialysis for at least 3 months. A significant relation was evident between anemia and LV hypertrophy. The mean LV mass index was 158 +/- 6 g/m2 (mean +/- standard error) in patients in the lowest quartile of serum hemoglobin and 140 +/- 10, 132 +/- 7 and 120 +/- 8 g/m2 in the second, third and uppermost quartiles, respectively (p = 0.005). This relation persisted after adjusting for systolic blood pressure, treatment mode and suspected coronary artery disease. Forty-eight patients had paired studies. In these, LV mass index increased as hemoglobin decreased (coefficient = -0.81 g/m2/g/liter, p less than 0.025). These data indicate that anemia contributes to the development of LV hypertrophy in patients with end-stage renal disease.
Adult↗
Conformationally restricted analogues of atriopeptin(103-125)amide.
Conformationally restricted analogues of atriopeptin(103-125)amide were prepared by synthesizing novel bicyclic peptides in which a second disulfide bridge linking residues 108 and 117 was introduced. These syntheses were shown to proceed with no significant scrambling of the disulfide bonds and demonstrated that structurally defined bicyclic analogues of atrial peptides could be easily prepared. The conformationally restrained analogues described here were found to be biologically active with potencies (EC50s) ranging from 0.05 to 3 microM. In addition, these bicyclic peptides (and many of the monocyclic precursors) were found to bind selectively to a class of specific tissue binding sites that have not been shown to be associated with any known second messenger system (NVR binding sites). Since affinity for the receptor class linked to vasorelaxation was negatively affected by the conformational restrictions described here, binding of atrial peptides to this class of receptors appears to have more specific conformational requirements than does binding to the NVR sites.
Amino Acid Sequence↗
Identification of structural requirements for analogues of atrial natriuretic peptide (ANP) to discriminate between ANP receptor subtypes.
The structure-activity relationships for affinity and selective binding of atrial natriuretic peptide (ANP) and analogues to guanylate cyclase coupled (CC) and non-cyclase coupled (NC) receptors in rabbit lung membranes are described. We have designed a series of peptides to try to identify the minimal sequence involved in specific recognition of each receptor subtype. The affinity of the peptides was determined from competitive binding experiments. Several peptides derived from the rat ANP sequence, e.g., des-[Phe106, Gly107, Ala115, Gln116]ANP-(103-125)NH2 (4), des-[Cys105,121]ANP-(104-126) (5), and [Acm-Cys105]ANP-(105-114)NH2 (9) have high affinity and selectivity for the noncoupled site. Peptide 4 was the most selective ligand with an affinity superior to that of ANP-(103-126). This compound does not displace the radiolabeled ligand from the guanylate cyclase coupled receptor at the highest concentration tested (100 nM). The structure-activity relationship for affinity and selectivity is discussed. Comparison of the peptide sequences suggests that the structural feature responsible for recognition of the NC site resides in a single sequence of seven contiguous amino acids from the cyclic core of the hormone. The corresponding heptapeptide retains affinity to the guanylate cyclase uncoupled binding site and is proposed to encompass the minimal sequence for specific recognition of the non-guanylate cyclase coupled ANP receptor.
Amino Acid Sequence↗
Analogues of atriopeptin(103-125)amide having high binding selectivity.
Analogues of atriopeptin(103-125)amide were prepared having a disulfide bridge at positions different from that found in the natural product. Most of these conformationally perturbed peptides were found to bind selectively to one subclass of binding sites. Binding affinity to a class of specific binding sites that is not associated with any known biological activity (nonvasorelaxant or NVR binding sites) is unaffected or even modestly improved. Affinity for the receptor subclass that is associated with vasorelaxation (VR subclass) decreases in most examples. In several cases, binding to the VR subclass is below the limits of detection for the assay used here. The data demonstrate that binding of atrial peptides to VR receptors requires rigidly defined receptor/ligand interactions. In contrast, the NVR subclass of binding sites appears to tolerate changes in peptide structure quite well.
Animals↗
A linear analog of atrial natriuretic peptide (ANP) discriminates guanylate cyclase-coupled ANP receptors from non-coupled receptors.
Atrial natriuretic peptide (ANP) contains a disulfide which is generally considered to be required for biological activity. A truncated linear ANP analog, des-Cys105,Cys121-ANP-(104-126) (referred to as analog I), that lacks the 2 cysteine residues of the parent peptide was synthesized. In competition binding studies using rabbit lung membranes, ANP-(103-126) and analog I displaced bound 125I-ANP-(103-126) from specific ANP binding sites 100 and 73%, respectively. The concentrations of ANP-(103-126) and analog I that produced 50% inhibition of radioligand binding to the membranes were 0.26 +/- 0.07 and 0.31 +/- 0.09 nM, respectively. Radioiodinated ANP-(103-126) and analog I were chemically cross-linked to binding sites on rabbit lung membranes, and the labeled membrane proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. 125I-Analog I specifically labeled a 65,000-dalton protein and a 135,000-dalton protein which, under reducing conditions, dissociated into 65,000-dalton subunits. In contrast, 125I-ANP-(103-126) labeled specifically a nonreducible 135,000-dalton protein, in addition to the 65,000-dalton species and the reducible 135,000-dalton species. ANP-(103-126) (100 nM) stimulated rabbit lung particulate guanylate cyclase activity, whereas analog I, at the same concentration, had no effect on cyclic GMP production and did not antagonize the effect of ANP-(103-126). From these observations, we conclude that analog I is a selective ligand which binds to approximately 73% of the total ANP binding sites present in rabbit lung membranes. Unlike ANP-(103-126), analog I does not bind to the remaining 27% of the binding sites and does not activate guanylate cyclase. Binding to the cyclase-linked ANP receptor correlates with the specific labeling by 125I-ANP-(103-126) of the nonreducible 135,000-dalton membrane protein.
Animals↗
Phosphorylation of high- and low-molecular-mass atrial natriuretic peptide analogs by cyclic AMP-dependent protein kinase.
Synthetic high- and low-molecular-mass atrial peptides were phosphorylated in vitro by cyclic AMP-dependent protein kinase and [32P]ATP. From a series of atrial peptide analogs, it was deduced that the amino acid sequence, Arg101-Ser104 of atriopeptin was required for optimal phosphorylation. Phosphorylated AP(99-126) was less potent than the parent atriopeptin in vasorelaxant activity and receptor-binding properties. These results indicate that the presence of a phosphate group at the N-terminus of AP(99-126) decreases the interaction of the peptide with its receptor and, as a consequence, decreases bioactivity. These observations are in contrast to those of Rittenhouse et al. [(1986) J. Biol. Chem. 261, 7607-7610] who reported that phosphorylation of AP(101-126) enhanced the stimulation of Na/K/Cl cotransport in cultured vascular smooth muscle cells.
Animals↗