Biomedical subjects

D S King

Publications and source records attributed to D S King.

At least 19 recordsLinked to original sources

PubMed2001

Molecular determinants of receptor binding and signaling by the CX3C chemokine fractalkine.

Fractalkine/CX3CL1 is a membrane-tethered chemokine that functions as a chemoattractant and adhesion protein by interacting with the receptor CX3CR1. To understand the molecular basis for the interaction, an extensive mutagenesis study of fractalkine's chemokine domain was undertaken. The results reveal a cluster of basic residues (Lys-8, Lys-15, Lys-37, Arg-45, and Arg-48) and one aromatic (Phe-50) that are critical for binding and/or signaling. The mutant R48A could bind but not induce chemotaxis, demonstrating that Arg-48 is a signaling trigger. This result also shows that signaling residues are not confined to chemokine N termini, as generally thought. F50A showed no detectable binding, underscoring its importance to the stability of the complex. K15A displayed unique signaling characteristics, eliciting a wild-type calcium flux but minimal chemotaxis, suggesting that this mutant can activate some, but not all, pathways required for migration. Fractalkine also binds the human cytomegalovirus receptor US28, and analysis of the mutants indicates that US28 recognizes many of the same epitopes of fractalkine as CX3CR1. Comparison of the binding surfaces of fractalkine and the CC chemokine MCP-1 reveals structural details that may account for their dual recognition by US28 and their selective recognition by host receptors.

Amino Acid Sequence↗

PubMed2001

Multi-site phosphorylation of Pho4 by the cyclin-CDK Pho80-Pho85 is semi-processive with site preference.

As part of a nutrient-responsive signaling pathway, the budding yeast cyclin-CDK complex Pho80-Pho85 phosphorylates the transcription factor Pho4 on five sites and inactivates it. Here, we describe the kinetic reaction between Pho80-Pho85 and Pho4. Through experimentation and computer modeling we have determined that Pho80-Pho85 phosphorylates Pho4 in a semi-processive fashion that results from a balance between kcat and k(off). In addition, we show that Pho80-Pho85 phosphorylates certain sites preferentially. Phosphorylation of the site with the highest preference inhibits the transcriptional activity of Pho4 when it is in the nucleus, while phosphorylation of the lowest-preference sites is required for export of Pho4 from the nucleus. This method of phosphorylation may allow Pho80-Pho85 to quickly inactivate Pho4 in the nucleus and efficiently phosphorylate Pho4 to completion.

Binding Sites↗

PubMed2001

Treatment of methicillin-resistant Staphylococcus epidermidis infection following repair of an ulnar fracture and humeroradial joint luxation in a horse.

A 27-month-old Rocky Mountain Horse was examined because of a fracture of the proximal portion of the ulna and luxation of the humeroradial joint (Monteggia fracture). Open reduction was performed, using a mechanical distractor, and the ulnar fracture was stabilized by application of a bone plate and screws. After surgery, the horse developed an infection of the surgical site, and bacterial culture of fluid from the surgical site yielded a pure growth of methicillin-resistant Staphylococcus epidermidis susceptible to oxytetracycline, erythromycin, rifampin, and vancomycin. Treatment with oxytetracycline did not result in a favorable clinical response. Therefore, the horse was treated systemically with vancomycin and rifampin, and vancomycin-impregnated polymethyl methacrylate beads were implanted at the surgical site. Six months after surgery, the horse was sound at a walk or trot, and bony union was evident on radiographs of the elbow joint.

Animals↗

PubMed2001

Effects of androstenedione-herbal supplementation on serum sex hormone concentrations in 30- to 59-year-old men.

The effectiveness of a nutritional supplement designed to enhance serum testosterone concentrations and prevent the formation of dihydrotestosterone and estrogens from the ingested androgens was investigated in healthy 30- to 59-year old men. Subjects were randomly assigned to consume DION (300 mg androstenedione, 150 mg dehydroepiandrosterone, 540 mg saw palmetto, 300 mg indole-3-carbinol, 625 mg chrysin, and 750 mg Tribulus terrestris per day; n = 28) or placebo (n = 27) for 28 days. Serum free testosterone, total testosterone, androstenedione, dihydrotestosterone, estradiol, prostate-specific antigen (PSA), and lipid concentrations were measured before and throughout the 4-week supplementation period. Serum concentrations of total testosterone and PSA were unchanged by supplementation. DION increased (p < 0.05) serum androstenedione (342%), free testosterone (38%), dihydrotestosterone (71%), and estradiol (103%) concentrations. Serum HDL-C concentrations were reduced by 5.0 mg/dL in DION (p < 0.05). Increases in serum free testosterone (r2 = 0.01), androstenedione (r2 = 0.01), dihydrotestosterone (r2 = 0.03), or estradiol (r2 = 0.07) concentrations in DION were not related to age. While the ingestion of androstenedione combined with herbal products increased serum free testosterone concentrations in older men, these herbal products did not prevent the conversion of ingested androstenedione to estradiol and dihydrotestosterone.

Adjuvants, Immunologic↗

PubMed2001

Engineering the prion protein using chemical synthesis.

In recent years, the technology of solid-phase peptide synthesis (SPPS) has improved to the extent that chemical synthesis of small proteins may be a viable complementary strategy to recombinant expression. We have prepared several modified and wild-type prion protein (PrP) polypeptides, of up to 112 residues, that demonstrate the flexibility of a chemical approach to protein synthesis. The principal event in prion disease is the conformational change of the normal, alpha-helical cellular protein (PrPc) into a beta-sheet-rich pathogenic isoform (PrP(Sc)). The ability to form PrP(Sc) in transgenic mice is retained by a 106 residue 'mini-prion' (PrP106), with the deletions 23-88 and 141-176. Synthetic PrP106 (sPrP106) and a His-tagged analog (sPrP106HT) have been prepared successfully using a highly optimized Fmoc chemical methodology involving DCC/HOBt activation and an efficient capping procedure with N-(2-chlorobenzyloxycarbonyloxy) succinimide. A single reversed-phase purification step gave homogeneous protein, in excellent yield. With respect to its conformational and aggregational properties and its response to proteinase digestion, sPrP106 was indistinguishable from its recombinant analog (rPrP106). Certain sequences that proved to be more difficult to synthesize using the Fmoc approach, such as bovine (Bo) PrP(90-200), were successfully prepared using a combination of the highly activated coupling reagent HATU and t-Boc chemistry. To mimic the glycosylphosphatidyl inositol (GPI) anchor and target sPrP to cholesterol-rich domains on the cell surface, where the conversion of PrPc is believed to occur, a lipophilic group or biotin, was added to an orthogonally side-chain-protected Lys residue at the C-terminus of sPrP sequences. These groups enabled sPrP to be immobilized on either the cell surface or a streptavidin-coated ELISA plate, respectively, in an orientation analogous to that of membrane-bound, GPI-anchored PrPc. The chemical manipulation of such biologically relevant forms of PrP by the introduction of point mutations or groups that mimic post-translational modifications should enhance our understanding of the processes that cause prion diseases and may lead to the chemical synthesis of an infectious agent.

Amino Acid Sequence↗

PubMed2001

Crystal structure of an activated response regulator bound to its target.

The chemotactic regulator CheY controls the direction of flagellar rotation in Escherichia coli. We have determined the crystal structure of BeF3--activated CheY from E. coli in complex with an N-terminal peptide derived from its target, FliM. The structure reveals that the first seven residues of the peptide pack against the beta4-H4 loop and helix H4 of CheY in an extended conformation, whereas residues 8-15 form two turns of helix and pack against the H4-beta5-H5 face. The peptide binds the only region of CheY that undergoes noticeable conformational change upon activation and would most likely be sandwiched between activated CheY and the remainder of FliM to reverse the direction of flagellar rotation.

Amino Acid Sequence↗

PubMed2001

Effects of resistance training on cardiovascular responses to lower body negative pressure in the elderly.

The purpose of the present study was to determine whether resistance training alters the cardiovascular responses to submaximal lower body negative pressure (LBNP) in the elderly. Twenty-one subjects were randomized into a control (C: n=10; 70 +/- 3 years, mean +/- SD) or a resistance training (TR: n=11; 67 +/- 7 years) group. Subjects in the TR underwent 12 weeks of training consisting of three sets of 8-12 contractions at approximately 60-80% of their initial maximal one repetition, three times per week, on 10 different machines. Before (Pre) and after (Post) training, all subjects underwent exposures of LBNP of -10, -20 and -40 Torr and muscle biopsy sampling at the vastus lateralis. TR increased (P< or =0.05) knee extension (Pre=379 +/- 140 N, Post=534 +/- 182 N) and chest press (Pre=349 +/- 137 N, Post=480 +/- 192 N) strength. Neither body weight nor percentage body fat were altered (P >0.05) by training. Resistance training increased (P< or =0.05) cross-sectional area in both Type I (4203 +/- 1196 to 5248 +/- 1728 microm2) and Type II (3375 +/- 1027 to 4286 +/- 1892 microm2) muscle fibres. Forearm blood flow, forearm vascular conductance, mean arterial pressure, and heart-rate responses to LBNP were not altered by the training. These data suggest that the cardiovascular responses of elderly to LBNP are unaffected by 12 weeks of whole-body resistance training despite increases in muscle strength and size.

Aged↗

PubMed2001

Endocrine and lipid responses to chronic androstenediol-herbal supplementation in 30 to 58 year old men.

OBJECTIVE: The effectiveness of an androgenic nutritional supplement designed to enhance serum testosterone concentrations and prevent the formation of dihydrotestosterone and estrogen was investigated in healthy 3 to 58 year old men. DESIGN: Subjects were randomly assigned to consume a nutritional supplement (AND-HB) containing 300-mg androstenediol, 480-mg saw palmetto, 450-mg indole-3-carbinol, 300-mg chrysin, 1,500 mg gamma-linolenic acid and 1.350-mg Tribulus terrestris per day (n = 28), or placebo (n = 27) for 28 days. Subjects were stratified into age groups to represent the fourth (30 year olds, n = 20), fifth (40 year olds, n = 20) and sixth (50 year olds, n = 16) decades of life. MEASUREMENTS: Serum free testosterone, total testosterone, androstenedione, dihydrotestosterone, estradiol, prostate specific antigen and lipid concentrations were measured before supplementation and weekly for four weeks. RESULTS: Basal serum total testosterone, estradiol, and prostate specific antigen (PSA) concentrations were not different between age groups. Basal serum free testosterone concentrations were higher (p < 0.05) in the 30- (70.5 +/- 3.6 pmol/L) than in the 50 year olds (50.8 +/- 4.5 pmol/L). Basal serum androstenedione and dihydrotestosterone (DHT) concentrations were significantly higher in the 30- (for androstenedione and DHT, respectively, 10.4 +/- 0.6 nmol/L and 2198.2 +/- 166.5 pmol/L) than in the 40- (6.8 +/- 0.5 nmol/L and 1736.8 +/- 152.0 pmol/L) or 50 year olds (6.0 +/- 0.7 nmol/L and 1983.7 +/- 147.8 pmol/L). Basal serum hormone concentrations did not differ between the treatment groups. Serum concentrations of total testosterone and PSA were unchanged by supplementation. Ingestion of AND-HB resulted in increased (p < 0.05) serum androstenedione (174%), free testosterone (37%), DHT (57%) and estradiol (86%) throughout the four weeks. There was no relationship between the increases in serum free testosterone, androstenedione, DHT, or estradiol and age (r2 = 0.08, 0.03, 0.05 and 0.02, respectively). Serum HDL-C concentrations were reduced (p < 0.05) by 0.14 mmol/L in AND-HB. CONCLUSIONS: These data indicate that ingestion of androstenediol combined with herbal products does not prevent the formation of estradiol and dihydrotestosterone.

Administration, Oral↗

PubMed2001

Random circular permutation leading to chain disruption within and near alpha helices in the catalytic chains of aspartate transcarbamoylase: effects on assembly, stability, and function.

A collection of circularly permuted catalytic chains of aspartate transcarbamoylase (ATCase) has been generated by random circular permutation of the pyrB gene. From the library of ATCases containing permuted polypeptide chains, we have chosen for further investigation nine ATCase variants whose catalytic chains have termini located within or close to an alpha helix. All of the variants fold and assemble into dodecameric holoenzymes with similar sedimentation coefficients and slightly reduced thermal stabilities. Those variants disrupted within three different helical regions in the wild-type structure show no detectable enzyme activity and no apparent binding of the bisubstrate analog N:-phosphonacetyl-L-aspartate. In contrast, two variants whose termini are just within or adjacent to other alpha helices are catalytically active and allosteric. As expected, helical disruptions are more destabilizing than loop disruptions. Nonetheless, some catalytic chains lacking continuity within helical regions can assemble into stable holoenzymes comprising six catalytic and six regulatory chains. For seven of the variants, continuity within the helices in the catalytic chains is important for enzyme activity but not necessary for proper folding, assembly, and stability of the holoenzyme.

Amino Acid Sequence↗

PubMed2001

Effect of prior exercise on glucose metabolism in trained men.

Several studies have demonstrated that oral glucose tolerance is impaired in the immediate postexercise period. A double-tracer technique was used to examine glucose kinetics during a 2-h oral glucose (75 g) tolerance test (OGTT) 30 min after exercise (Ex, 55 min at 71 +/- 2% of peak O(2) uptake) and 24 h after exercise (Rest) in endurance-trained men. The area under the plasma glucose curve was 71% greater in Ex than in Rest (P = 0.01). The higher glucose response occurred even though whole body rate of glucose disappearance was 24% higher after exercise (P = 0.04, main effect). Whole body rate of glucose appearance was 25% higher after exercise (P = 0.03, main effect). There were no differences in total (2 h) endogenous glucose appearance (R(a)E) or the magnitude of suppression of R(a)E, although R(a)E was higher from 15 to 30 min during the OGTT in Ex. However, the cumulative appearance of oral glucose was 30% higher in Ex (P = 0.03, main effect). There were no differences in glucose clearance rate or plasma insulin responses between the two conditions. These results suggest that adaptations in splanchnic tissues by prior exercise facilitate greater glucose output from the splanchnic region after glucose ingestion, resulting in a greater glycemic response and, consequently, a greater rate of whole body glucose uptake.

Adult↗

PubMed2000

Cyclooxygenase-2 enzyme inhibitors: place in therapy.

Nonsteroidal anti-inflammatory drugs (NSAIDs) play a major role in the management of inflammation and pain caused by arthritis. A new class of NSAIDs that selectively inhibit the cyclooxygenase-2 (COX-2) enzyme has been developed. The first COX-2 inhibitors, celecoxib and rofecoxib, are said to provide therapeutic benefit with less toxicity than traditional NSAIDs. A third COX-2-selective inhibitor, meloxicam, has recently been introduced. COX-2 inhibitors and traditional NSAIDs do not appear to differ significantly in their effectiveness in alleviating pain or inflammation. They have similar gastrointestinal side effects, including abdominal pain, dyspepsia and diarrhea. However, short-term studies show fewer gastrointestinal ulcers in patients treated with COX-2 inhibitors compared with traditional NSAIDs.

Analgesics, Non-Narcotic↗

PubMed2000

Structure and function of a human TAFII250 double bromodomain module.

TFIID is a large multiprotein complex that initiates assembly of the transcription machinery. It is unclear how TFIID recognizes promoters in vivo when templates are nucleosome-bound. Here, it is shown that TAFII250, the largest subunit of TFIID, contains two tandem bromodomain modules that bind selectively to multiply acetylated histone H4 peptides. The 2.1 angstrom crystal structure of the double bromodomain reveals two side-by-side, four-helix bundles with a highly polarized surface charge distribution. Each bundle contains an Nepsilon-acetyllysine binding pocket at its center, which results in a structure ideally suited for recognition of diacetylated histone H4 tails. Thus, TFIID may be targeted to specific chromatin-bound promoters and may play a role in chromatin recognition.

Acetylation↗

PubMed2000

Effects of anabolic precursors on serum testosterone concentrations and adaptations to resistance training in young men.

The effects of androgen precursors, combined with herbal extracts designed to enhance testosterone formation and reduce conversion of androgens to estrogens was studied in young men. Subjects performed 3 days of resistance training per week for 8 weeks. Each day during Weeks 1, 2, 4, 5, 7, and 8, subjects consumed either placebo (PL; n = 10) or a supplement (ANDRO-6; n = 10), which contained daily doses of 300 mg androstenedione, 150 mg DHEA, 750 mg Tribulus terrestris, 625 mg Chrysin, 300 mg Indole-3-carbinol, and 540 mg Saw palmetto. Serum androstenedione concentrations were higher in ANDRO-6 after 2, 5, and 8 weeks (p <.05), while serum concentrations of free and total testosterone were unchanged in both groups. Serum estradiol was elevated at Weeks 2, 5, and 8 in ANDRO-6 (p <.05), and serum estrone was elevated at Weeks 5 and 8 (p <.05). Muscle strength increased (p <.05) similarly from Weeks 0 to 4, and again from Weeks 4 to 8 in both treatment groups. The acute effect of one third of the daily dose of ANDRO-6 and PL was studied in 10 men (23 +/- 4 years). Serum androstenedione concentrations were elevated (p <.05) in ANDRO-6 from 150 to 360 min after ingestion, while serum free or total testosterone concentrations were unchanged. These data provide evidence that the addition of these herbal extracts to androstenedione does not result in increased serum testosterone concentrations, reduce the estrogenic effect of androstenedione, and does not augment the adaptations to resistance training.

Adaptation, Physiological↗

PubMed2000

Endocrine responses to chronic androstenedione intake in 30- to 56-year-old men.

In young men, chronic ingestion of 100 mg androstenedione (ASD), three times per day, does not increase serum total testosterone but does increase serum estrogen and ASD concentrations. We investigated the effects of ASD ingestion in healthy 30- to 56-yr-old men. In a double-blind, randomly assigned manner, subjects consumed 100 mg ASD three times daily (n = 28), or placebo (n = 27) for 28 days. Serum ASD, dihydrotestosterone (DHT), free and total testosterone, estradiol, prostate-specific antigen (PSA), and lipid concentrations were measured at week 0 and each week throughout the supplementation period. Serum total testosterone and PSA concentrations did not change with supplementation. Elevated serum concentrations of ASD (300%), free testosterone (45%), DHT (83%), and estradiol (68%) were observed during weeks 1-4 in ASD (P < 0.05). There was no relationship between age and changes in serum ASD (r2 = 0.024), free testosterone (r2 = 0.00), or estradiol (r2 = 0.029) concentrations with ASD, whereas the serum DHT response to ASD ingestion was related to age (r2 = 0.244; P < 0.05). Serum concentrations of high-density lipoprotein cholesterol were decreased by 10% during the supplementation period (P < 0.05). These results suggest that the ingestion of 100 mg ASD, three times per day, does not increase serum total testosterone or PSA concentrations but does elicit increases in ASD, free testosterone, estradiol, and DHT and decreases serum high-density lipoprotein cholesterol concentrations.

Administration, Oral↗

PubMed1999

A chemically synthesized version of the insect antibacterial glycopeptide, diptericin, disrupts bacterial membrane integrity.

Insects protect themselves against bacterial infection by secreting a battery of antimicrobial peptides into the hemolymph. Despite recent progress, important mechanistic questions, such as the precise bacterial targets, the nature of any cooperation that occurs between peptides, and the purpose of multiple peptide isoforms, remain largely unanswered. We report herein the chemical synthesis and preliminary mechanistic investigation of diptericin, an 82 residue glycopeptide that contains regions similar to two different types of antibacterial peptides. A revised, highly practical synthesis of the precursor N(alpha)-Fmoc-Thr(Ac(3)-alpha-D-GalNAc) allowed us to produce sufficient quantities of the glycopeptide for mechanistic assays. The synthetic, full-length polypeptide proved to be active in growth inhibition assays with an IC(50) of approximately 250 nM, a concentration similar to that found in the insect hemolymph. Biological analysis of diptericin fragments indicated that the main determinant of antibacterial activity lay in the C-terminal region that is similar to the attacin peptides, although the N-terminal segment, related to the proline-rich family of antibacterial peptides, augmented that activity by 100-fold. In all assays, activity appeared glycosylation independent. Circular dichroism of unglycosylated diptericin indicated that the peptide lacked structure both in plain buffer and in the presence of liposomes. Diptericin increased the permeability of the outer and inner membranes of Escherichia coli D22 cells, suggesting possible mechanisms of action. The ability to access glycopeptides of this type through chemical synthesis will facilitate further mechanistic studies.

Amino Acid Sequence↗

PubMed1999

Effect of oral androstenedione on serum testosterone and adaptations to resistance training in young men: a randomized controlled trial.

CONTEXT: Androstenedione, a precursor to testosterone, is marketed to increase blood testosterone concentrations as a natural alternative to anabolic steroid use. However, whether androstenedione actually increases blood testosterone levels or produces anabolic androgenic effects is not known. OBJECTIVES: To determine if short- and long-term oral androstenedione supplementation in men increases serum testosterone levels and skeletal muscle fiber size and strength and to examine its effect on blood lipids and markers of liver function. DESIGN AND SETTING: Eight-week randomized controlled trial conducted between February and June 1998. PARTICIPANTS: Thirty healthy, normotestosterogenic men (aged 19-29 years) not taking any nutritional supplements or androgenic-anabolic steroids or engaged in resistance training. INTERVENTIONS: Twenty subjects performed 8 weeks of whole-body resistance training. During weeks 1, 2, 4, 5, 7, and 8, the men were randomized to either androstenedione, 300 mg/d (n = 10), or placebo (n = 10). The effect of a single 100-mg androstenedione dose on serum testosterone and estrogen concentrations was determined in 10 men. MAIN OUTCOME MEASURES: Changes in serum testosterone and estrogen concentrations, muscle strength, muscle fiber cross-sectional area, body composition, blood lipids, and liver transaminase activities based on assessments before and after short- and long-term androstenedione administration. RESULTS: Serum free and total testosterone concentrations were not affected by short- or long-term androstenedione administration. Serum estradiol concentration (mean [SEM]) was higher (P<.05) in the androstenedione group after 2 (310 [20] pmol/L), 5 (300 [30] pmol/L), and 8 (280 [20] pmol/L) weeks compared with presupplementation values (220 [20] pmol/L). The serum estrone concentration was significantly higher (P<.05) after 2 (153 [12] pmol/L) and 5 (142 [15] pmol/L) weeks of androstenedione supplementation compared with baseline (106 [11] pmol/L). Knee extension strength increased significantly (P<.05) and similarly in the placebo (770 [55] N vs 1095 [52] N) and androstenedione (717 [46] N vs 1024 [57] N) groups. The increase of the mean cross-sectional area of type 2 muscle fibers was also similar in androstenedione (4703 [471] vs 5307 [604] mm2; P<.05) and placebo (5271 [485] vs 5728 [451] mm2; P<.05) groups. The significant (P<.05) increases in lean body mass and decreases in fat mass were also not different in the androstenedione and placebo groups. In the androstenedione group, the serum high-density lipoprotein cholesterol concentration was reduced after 2 weeks (1.09 [0.08] mmol/L [42 (3) mg/dL] vs 0.96 [0.08] mmol/L [37 (3) mg/dL]; P<.05) and remained low after 5 and 8 weeks of training and supplementation. CONCLUSIONS: Androstenedione supplementation does not increase serum testosterone concentrations or enhance skeletal muscle adaptations to resistance training in normotestosterogenic young men and may result in adverse health consequences.

Administration, Oral↗

PubMed1999

Biochemical and biophysical characterization of the trimerization domain from the heat shock transcription factor.

Previously, we had characterized a 91 amino acid fragment of the heat shock transcription factor from the yeast Kluyveromyces lactis and had shown it to be highly alpha-helical and sufficient for formation of homotrimers [Peteranderl, R., and Nelson, H. C. M. (1992) Biochemistry 31, 12272-12276]. Based on those data, as well as the presence of hydrophobic heptad repeats, we postulated that the trimerization domain contains a three-stranded coiled-coil and that it might resemble the trimerization domain found in influenza hemagglutinin. Here, we further characterize the trimerization domain and show that the minimal domain needs 71 residues to remain trimeric and highly alpha-helical. 19F NMR spectroscopy suggests that the structure contains three parallel strands that are in register along the long axis of the coiled-coil. Electron paramagnetic resonance spectroscopy studies show that the C-termini of the subunits are in close proximity; this is in contrast to the topology of the hemaglutinin trimerization domain where the C-termini form buttressing helices. Analytical ultracentrifugation also confirms that the structure is elongated and unlikely to have buttressing helices. Additional experiments suggest that the trimerization domain has at least two subdomains. The first subdomain has the potential to form trimers independently, though not as stably as the complete domain. The second subdomain is quite helical, forms large oligomers, and appears to provide stability to the complete domain. Our current model for the heat shock transcription factor trimerization domain is a highly elongated coiled-coil structure, with a potential break in the coiled-coil region located between the two subdomains.

Amino Acid Sequence↗

PubMed1999

Biological actions of synthetic locust ion transport peptide (ITP).

Locust Ion Transport Peptide (ITP) a member of the arthropod neuropeptide family which includes hyperglycemic, vitellogenesis-inhibiting, and moult-inhibiting hormones (CHH, VIH, MIH, respectively) was synthesized as proposed by Meredith et al. (1996) with terminal amidation of amino acid residue 72 and with 3 disulphide bridges. This is the first member of this family to be synthesized. Biological activities of synthetic ITP (synITP) were very similar to those previously reported for ITP purified from Schistocerca corpora cardiaca (ScgITP) and partially sequenced by Audsley et al. (1992a, b). Dose-response curves for both synITP and ScgITP on ileal transport of Cl- (measured as increased short-circuit current, delta Isc), were similar with a EC50 of 1-2 nM. The Isc time course and maximum delta Isc across ileal epithelia at different dosages of synITP and ScgITP had similar patterns as did changes in transepithelial open-circuit potential (Vt) and resistance (Rt), reflecting changes in salt transport which drives fluid absorption. Disulphide bridges were shown to be required for biological activity of synITP, which caused the same 4-fold increase in ileal fluid transport rate (Jv) as previously reported for ScgITP. Both synITP and ScgITP caused only partial stimulation of rectal Isc and had no significant effect on rectal Jv. These results indicate that the structure of ITP predicted earlier from cDNA is correct.

Animals↗