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PubMed1989

Effect of the interaction of drug-beta-cyclodextrin complex with bile salts on the drug absorption from rat small intestinal lumen.

This investigation was concerned with the change of the bioavailability of a drug owing to the interaction of the drug-beta-cyclodextrin complex with bile salts in rat intestinal lumen. The absorption of sulfamethizole (SMZ) from rat intestinal lumen after administration of SMZ-beta-cyclodextrin complex was determined by a closed-loop method in the presence or absence of bile. The blood level of SMZ after administration of SMZ-beta-cyclodextrin complex was significantly decreased in comparison with that after administration of SMZ alone in bile duct-ligated rats. On the other hand, the blood level of SMZ after SMZ-beta-cyclodextrin administration in intact rats (bile duct non-ligated) or on the addition of sodium cholate was similar to the level in the case of SMZ alone. Thus, bile salts were found to act as a competing agent in the gastrointestinal tract.

Animals↗

PubMed1994

Dissolution behavior of phenytoin-bile salt complexes prepared by co-grinding.

The physicochemical properties of phenytoin (PHT)-bile salt complexes comprised of sodium dehydrocholate (DHCNa), sodium deoxycholate (DCNa) or sodium cholate (CNa) prepared by co-grinding were investigated by X-ray diffraction analysis, DSC and dissolution kinetics. All X-ray diffraction peak intensities of the co-ground PHT-bile salt [1 mol: 1 mol] mixtures were decreased by grinding for 3 h, and showed a halo pattern of a noncrystalline solid. The solubility of ground products with DCNa, DHCNa and CNa were 212, 56, 68 times higher, respectively, than those of physical mixtures.

Bile Acids and Salts↗

PubMed1990

The interaction of acetylcholine receptors in porcine atrial membranes with three kinds of G proteins.

We developed a simple procedure to detect the interaction of muscarinic receptors in atrial membranes with exogenous GTP-binding proteins (G proteins). The procedure consists of mixing atrial membranes with G proteins in the presence of sodium cholate, diluting the mixture with a salt buffer and then measuring the ligand binding activity. The displacement by carbachol of [3H] QNB binding to muscarinic receptors in the atrial membranes was not affected by guanine nucleotides when the membranes had been treated at 60 degrees C for 30 min or with N-ethylmeleimide (NEM) and became affected by them after mixing the heat- or NEM-treated membranes with G proteins. The displacement curves in the presence of GTP were essentially the same irrespective of the presence or absence of G proteins. Those in the absence of GTP shifted to a lower concentration of carbachol with addition of a higher concentration of G proteins, indicating an increase in GTP-sensitive high affinity agonist binding sites. The highest affinity for carbachol was detected with membranes treated with NEM and then mixed with G proteins. The GTP-sensitive high affinity agonist binding could be detected with any one of three kinds of G proteins (Gi, Go, Gn) which were purified from porcine cerebrum, indicating that the muscarinic receptor m2 subtype may interact with and possibly activate these three kinds of G proteins.

Animals↗

PubMed1994

A study of the biological pharmacology of IFO, a new selective and reversible monoamine oxidase-B inhibitor.

3-[4-[3-(1H-Imidazol-1-yl)propoxy]phenyl]-5-trifluoromethyl-1,2,4- oxadiazole (IFO), designed to be a novel selective inhibitor of monoamine oxidase (MAO), showed highly selective inhibition for type-B (MAO-B); its IC50 was approximately > 200 microM and 30 nM for type-A (MAO-A) and MAO-B, respectively, in the standard assay using mitochondrial preparations from rat brain or liver. The in vitro MAO-B inhibition by IFO was time-independent, non-competitive and tight-binding; and furthermore, in the presence of sodium cholate its inhibition was not tight-binding and was reversible. Oral administration of IFO (0.5-100 mg/kg) produced a dose-dependent MAO-B inhibition in mouse brain; its ED50 (p.o., 1 hr) was 1.6 mg/kg, while L-deprenyl inhibited the enzyme with the ED50 of approximately 8.0 mg/kg. The ED50 for MAO-A was > 100 mg/kg for either IFO and L-deprenyl. The MAO inhibitive effect of IFO in mouse liver was the same as that in the brain, but that of L-deprenyl in mouse liver was different from that in the brain as shown by the ED50 values of 35 mg/kg and 0.6 mg/kg for MAO-A and MAO-B, respectively. In mice, IFO increased the striatal concentrations of 2-phenylethylamine (2-PEA) and showed almost the same protective efficacy as L-deprenyl against the lethality and striatal dopamine (DA) depletion induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). These results indicate that IFO appears to be a potent inhibitor of MAO-B in mouse brain.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine↗

PubMed1996

Strain-specific response to hypercholesterolaemic diets in the rat.

The cholesterolaemic effect of 2 hypercholesterolaemic diets was tested in 12 rat inbred strains. Diet I is a commercial diet supplemented with 2.0% (w/w) cholesterol and 5.0% (w/w) olive oil; diet II is identical to diet I with addition of 0.5% (w/w) sodium cholate. Strains with the highest plasma cholesterol response after diet I (BN and LEW) also had the highest cholesterol response after diet II (hyperresponders, mean response > 3.5 mmol/l). In the strains DA, SHR, BC, WAG, LOU, PVG and BUF the strain mean cholesterol response remained below 1.3 mmol/l after both diets (hyporesponders). Strains F344 and OM had an intermediate cholesterol response after both diets (normoresponders, mean response between 1.3 and 3.5 mmol/l). Only in the strains LOU, PVG and SHR there appeared to be a significant higher cholesterol response after diet II when compared with the cholesterol response after diet I. In the strain WKY this difference was of a borderline significance (P = 0.052) and this strain turned from a normoresponder after diet I into a hyperresponder after diet II. Liver cholesterol levels as measured after feeding diet II for two weeks also appeared to be strain-specific. No correlation was found between the plasma cholesterol response after diet II and the liver cholesterol levels. Changes in plasma phospholipid and triglyceride levels have been measured for both diet I and diet II. For group means a correlation between the cholesterol response and the change in phospholipid levels was found (r = 0.86 for diet I, P < 0.001 and r = 0.76 for diet II, P < 0.01). No such correlation was found for triglyceride levels.

Animals↗

PubMed1999

A mixture of organisms affects cholesterol metabolism together with rat cecal flora.

The effects of a mixture of organisms on cecal fermentation and cholesterol metabolism in sham-operated and cecectomized rats were investigated. Male F344 rats, allocated into four groups: cecectomized rats fed a mixture of organisms (CEMO), cecectomized rats fed rice bran (CERB), sham-operated rats fed a mixture of organisms (SHMO), and sham-operated rats fed rice bran (SHRB) for 4 weeks. The diets had 0.5% cholesterol and 0.125% sodium cholate added. There were no significant differences in the body weight gain and food intake among the groups. The cecal pH in the SHMO group was significantly lower than that in the other groups. The total cholesterol and (VLDL + IDL + LDL)-cholesterol concentrations in serum were significantly lower in the SHMO group than that in the SHRB group, and the triacylglycerol concentration in the sham-operated rats tended to decrease compared to the cecectomized rats. The fecal cholesterol excretion in the CERB group was higher than that in the other groups, and that in the SHMO group was significantly higher than in the SHRB group. The acetic acid, propionic acid, n-butyric acid, and total short-chain fatty acid concentrations in the cecum contents were significantly higher in the SHMO group than those in the other groups. Streptococcus, Bifidobacterium, and Lactobacillus in the SHMO group tended to be higher than the other groups and Bacteroidaceae in the CEMO and CERB groups were significantly higher than that in the SHMO group. The results demonstrate that the mixture of organisms was fermented with the cecal contents and that the metabolites such as short-chain fatty acid lowered the serum total cholesterol and liver cholesterol concentrations in the rats fed a cholesterol-containing diet.

Animals↗

PubMedDate not supplied

Safety and pharmacokinetics of an endotoxin-binding phospholipid emulsion.

BACKGROUND: Lipids and lipoproteins have been shown to bind and neutralize endotoxin and to improve outcomes in animal models of sepsis. OBJECTIVE: To provide safety and pharmacokinetic data for a protein-free, phospholipid-rich emulsion developed as an agent to neutralize endotoxin, and to study the changes in lipids and lipoproteins following emulsion administration. METHODS: Thirty healthy male volunteers (aged 18-45 y) were given an emulsion containing 92.5% soy phospholipid, 7.5% soy triglyceride, and 18 mM sodium cholate using a double-blind, placebo-controlled crossover protocol. Emulsion at 3 escalating doses (75, 150, 300 mg/kg) based on phospholipid content was administered by intravenous infusion over 2 hours in the low- and mid-dose groups and 6 hours in the high-dose group. RESULTS: All subjects completed the protocol without significant toxicities. A slight dose-dependent increase in indirect bilirubin at the 24-hour time point was observed in the emulsion treatment period, with a maximum difference between placebo and emulsion of 0.9 mg/dL. Mean +/- SD peak phospholipid levels were 316 +/- 30, 533 +/- 53, and 709 +/- 86 mg/dL, and phospholipid half-lives were 5.4 +/- 0.6, 5.4 +/- 0.5, and 8.0 +/- 0.8 hours for the low, mid, and high doses, respectively. Increases in total cholesterol, low-density lipoprotein cholesterol and apolipoprotein A-I and B levels were observed. High-density lipoprotein cholesterol decreased immediately following emulsion infusion, but rebounded to above placebo levels by 24 hours. CONCLUSIONS: A unique phospholipid-rich emulsion was shown to have a favorable safety profile and to expand the blood lipid and lipoprotein pool without the use of human-derived blood products. Lipid levels expected to protect against the physiologic effects of bacterial endotoxin were achieved.

Adolescent↗

PubMed1991

Lysosomal and plasma membrane ganglioside GM3 sialidases of cultured human fibroblasts. Differentiation by detergents and inhibitors.

Cultured human fibroblasts contain two sialidases that degrade gangliosides such as GM3: a lysosomal activity that appears identical with the activity towards water-soluble substrates and that is deficient in the genetic lysosomal disorder sialidosis, and another enzyme that seems localized on the external surface of the plasma membrane. In this report we show that both enzymes can be differentiated in the presence of each other by choice of the detergent used for activation, and also by the inhibitory action of some polyanionic compounds such as sulphated glycosaminoglycans. The lysosomal ganglioside GM3 sialidase is greatly stimulated by sodium glycodeoxycholate and, to lesser degrees, by sodium glycocholate and sodium cholate. The ganglioside GM3 sialidase of the plasma membrane is not measurably active under the conditions of the lysosomal enzyme but is specifically activated by the non-ionic detergent Triton X-100. The glycodeoxycholate-stimulated, but not the Triton-activated, ganglioside GM3 sialidase activity was profoundly diminished in cell lines from patients with the lysosomal disorders sialidosis and galactosialidosis; however, both activities were normal in fibroblasts from patients with mucolipidosis IV, previously thought to be a ganglioside sialidase deficiency disorder. Both the lysosomal and the plasma membrane ganglioside GM3 sialidases were inhibited by sialic acids, suramin, dextran sulphate and sulphated glycosaminoglycans. Among the latter, heparin and heparan sulphate showed a much higher inhibitory potency towards the plasma membrane ganglioside GM3 sialidase than towards the lysosomal onw.(ABSTRACT TRUNCATED AT 250 WORDS)

Carbohydrate Metabolism, Inborn Errors↗

PubMedDate not supplied

Stimulation of pancreatic lipase activity by saponins isolated from Medicago sativa L.

Saponins isolated from Medicago sativa L. appeared to stimulate lipolytic activity and did not influence amylolytic or proteolytic activity of Neopancreatinum (extract of pancreatic enzymes: lipase, proteases and amylases). Saponins isolated from the aerial part of Medicago sativa L. were demonstrated to stimulate lipase activity more effectively than saponins separated from the root of this plant. When saponins were treated with 0.02 M HCl at 37 degrees C for 1 hour their lipase stimulation ability did not change. The saponins stimulated Neopancreatinum lipolytic activity more intensively in the presence of sodium cholate.

Amylases↗

PubMed2006

Transfersomes for transdermal drug delivery.

Transfersomes (Idea AG) are a form of elastic or deformable vesicle, which were first introduced in the early 1990s. Elasticity is generated by incorporation of an edge activator in the lipid bilayer structure. The original composition of these vesicles was soya phosphatidyl choline incorporating sodium cholate and a small concentration of ethanol. Transfersomes are applied in a non-occluded method to the skin and have been shown to permeate through the stratum corneum lipid lamellar regions as a result of the hydration or osmotic force in the skin. They have been used as drug carriers for a range of small molecules, peptides, proteins and vaccines, both in vitro and in vivo. It has been claimed by Idea AG that intact Transfersomes penetrate through the stratum corneum and the underlying viable skin into the blood circulation. However, this has not been substantiated by other research groups who have extensively probed the mechanism of penetration and interaction of elastic vesicles in the skin. Structural changes in the stratum corneum have been identified, and intact elastic vesicles visualised within the stratum corneum lipid lamellar regions, but no intact vesicles have been ascertained in the viable tissues. Using the principle of incorporating an edge-activator agent into a bilayer structure, a number of other elastic vesicle compositions have been evaluated. This review describes the research into the development and evaluation of Transfersomes and elastic vesicles as topical and transdermal delivery systems.

Administration, Cutaneous↗

PubMedDate not supplied

Effect of solvent polarity and viscosity on the guest binding dynamics with bile salt aggregates.

Bile salts form supramolecular aggregates with two binding sites with different properties. The guest binding dynamics to the aggregates and guest protection from species in the aqueous phase were investigated using fluorescence and laser flash photolysis experiments. Sodium cholate, deoxycholate and taurodeoxycholate were used as bile salts and acetonitrile or ethylene glycol were added as co-solvents to water in order to alter the binding properties of 1-ethylnaphthalene and 1-naphthyl-1-ethanol with the aggregates. The binding dynamics are faster and protection efficiencies are lower for guests bound to cholate and in the presence of either co-solvent.

Bile Acids and Salts↗

PubMed2005

Effect of Chlamydia pneumoniae infection and hyperlipidaemia on the expression of PPARgamma, P50 and c-Fos in aortic endothelial cells in C57bL/6J mice.

OBJECTIVE: To investigate the effect of C. pneumoniae infection and/or hyperlipidaemia on the expression of peroxisome proliferator-activated receptor gamma (PPARgamma), nuclear factor-kappa B (NF-kappaB) and activated protein-1 (AP-1) in aortic endothelial cells in C57BL/6J mice. METHODS AND RESULTS: Forty-eight, 8-week-old female C57BL/6J mice were divided into four groups:A, B, C and D (each twelve mice). Group A (as blank control) and B were fed a regular diet. Group C and D were fed an atherogenic diet (consisting of 15% fat, 2.5% cholesterol and 0.5% sodium cholate). Group B and D were infected with C. pneumoniae. Fourteen weeks later, the expression of PPARgamma, P50 (subunit of NF-kappaB) and c-Fos (subunit of AP-1) was determined by indirect immunofluorescence in the aortic endothelial cells. Slides of aortic sinus were prepared by cryosection, and stained with Sudan IV for examination of atherosclerotic plaque. The score of atherosclerotic plaque was determined by microscopy. The score of atherosclerotic plaque in group B was not increased, while it was significantly higher in groups C and D (P < 0.01), still the score in group D was higher than in group C (P < 0.01). The expression of PPARgamma, NF-kappaB and AP-1 in endothelial cells in aortic sinus was upregulated in group B, C and D, in comparison with group A (P < 0.05). There was no significant difference among groups B, C and D. CONCLUSION: The expression of PPARgamma, NF-kappaB and AP-1 was upregulated in the endothelial cells in mice infected with C. pneumoniae and/or fed with an atherogenic diet. An atherogenic diet or this diet combined with C. pneumoniae infection accelerated the process of atherosclerosis. The diet infected with C. pneumoniae alone would not accelerate this process. PPARgamma might play an anti-atherosclerotic role in this process.

Animals↗

PubMed2006

Isolation, characterization and study of enhancing effects on nasal absorption of insulin in rat of the total saponin from Acanthophyllum squarrosum.

OBJECTIVE: Isolation of the total saponins from Acanthophyllum squarrosum Boiss. and investigation of its surface activity, haemolytic effects on human erythrocytes as well as enhancing potentials on intranasal insulin absorption in rat in comparison with two other enhancers i.e. Quillaja total saponin (QTS) and sodium cholate (SC). MATERIALS AND METHODS: The decrease in blood glucose levels in five fasting rats following nasal administration of regular insulin solutions in the presence or absence of enhancers was determined by glucometric strips and used as an indication of insulin absorption. RESULTS: The results showed that ATS decreased surface tension of water to about 50 dyne.cm(-1) and caused complete haemolysis of human RBCs at a concentration of 250 microg.ml(-1). Following the instillation of solutions containing insulin and different absorption enhancers into the right nostril of rats, the percentage decrease in initial blood glucose was as follows: 72.46% (+/- 2.39%) for ATS, 63.22 % (+/-11.06%) for QTS and 60.06% (+/-14.93%) for SC. Percentage lowering in initial blood glucose concentrations against time showed that ATS exhibits a stronger effect than the two other enhancers although the difference was not statistically significant (p>0.05). CONCLUSION: ATS has a considerable absorption enhancing effect and can possibly be used to increase insulin bioavailability via nasal route. However the potential toxic effects of this saponin on nasal mucosa should be further evaluated.

Administration, Intranasal↗

PubMed1996

Renal effects of moderate hypercholesterolaemia in uninephrectomized rats.

Animal studies have shown that a 4-6-fold increase in serum cholesterol aggravates pre-existing renal injury. We studied the renal effects of moderate hypercholesterolaemia over a period of 18 weeks in uninephrectomized rats. Animals were allocated to two groups; the group 1 rats were fed a normal diet, as controls, and the group 2 rats were fed a high cholesterol diet containing 3% cholesterol and 1% sodium cholate by weight. The serum total cholesterol was higher in group 2 than in controls being 2.5 +/- 0.4 vs. 1.0 +/- 0.1 mmol l-1 at 9 weeks and 2.1 +/- 0.3 vs. 1.1 +/- 0.2 mmol l-1 at 18 weeks (p < 0.05 for both). Serum high density lipoprotein cholesterol levels were similar in both groups. The mean systolic blood pressure was higher in group 2 than in controls, at 145 +/- 9 vs. 137 +/- 8 mmHg (p < 0.05) by 13 weeks and 146 +/- 6 vs. 136 +/- 4 mmHg (p < 0.05) at 18 weeks. Serum creatinine and glomerular filtration rates were similar in both groups. Urine protein excretion remained within the normal range in both groups. Histological examination at 18 weeks showed diffuse fatty changes in the liver cells and prominent vacuolization of renal tubule cells in the group 2 rats. Nevertheless, the glomeruli were normal. There was no significant difference in mean glomerular volume between group 2 rats (1.20(-3) +/- 0.09(-3) mm3) and controls (1.36(-3) +/- 0.10(-3) mm3). Thus moderate hypercholesterolaemia for 18 weeks in uninephrectomized rats resulted in a mild elevation in blood pressure, but did not affect glomerular volume or glomerular histology, in spite of the deleterious effects on liver and renal tubule cells. We assume that extremely high levels of serum cholesterol are required to induce glomerulosclerosis in the rat.

Animals↗

PubMedDate not supplied

Polymer-coated liposomes: improved liposome stability and release of cytosine arabinoside (Ara-C).

Liposomes of dimyristoylphosphatidylcholine (DMPC) or dipalmitoylphosphatidylcholine (DPPC), cholesterol (CHOL) and dicetylphosphate (DCP) containing Ara-C were prepared at pH 4.5 and subsequently coated with a derivatized polysaccharide, O-palmitoylpullulan (OPP). Encapsulation efficiencies (EEF) of DMPC:CHOL:DCP (3:1:2 mol ratio) and DPPC:CHOL:DCP (3:1:0.5 mol ratio) liposomes were respectively 28 and 40% before and 15 and 27% after coating with polymer. Stabilities of liposomes in sodium cholate (SC) concentrations up to 16 mM at pH 5.6 or in SC solutions at pH 7.4 were increased by coating with OPP. At pH 2.0 and 37 degrees C, Ara-C was released at a slow rate, kuo (uncoated) or kco (coated), for 24 h after an initial rapid release phase that lasted for about 6 h and the ratio kuo/kco, was 1.9 and 5.7 for the DMPC:CHOL:DCP and the DPPC:CHOL:DCP compositions respectively. At pH 7.4 and 37 degrees C in the presence of 10 mM SC, the ratio kuo/kco was 5.1 and 1.4 respectively. It is concluded that coating liposomes with OPP increases the potential of using liposomes as a drug delivery system in harsh environments such as encountered after oral administration, although the magnitude of the effect is dependent on the liposome composition.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1987

Fractionation and reconstitution of the sarcoplasmic reticulum Ca2+ pump solubilized and stabilized by CHAPS/lipid micelles.

A procedure for solubilization, fractionation, and reconstitution of sarcoplasmic reticulum (SR) protein is presented. The SR protein is solubilized with the zwitterionic detergent CHAPS in the presence of added 5-mM phosphatidylcholine and 20% glycerol, which stabilize the reconstitutable Ca2+ transport activity. For reconstitution the solubilized SR protein is incorporated into preformed French-pressed unilamellar vesicles that had been treated with 10-mM sodium cholate. By passing the proteoliposomes through a centrifuged Sephadex G-50 column that had been equilibrated with potassium oxalate, the detergent is removed, and the proteoliposomes become sealed with potassium oxalate trapped inside. This procedure requires less than 2 h and results in Ca2+ uptake active of over 1 mumol/min/mg of protein. The solubilized SR protein was fractionated on a DEAE-Biogel A column. A fraction containing the Ca2+-ATPase but not the Mr 55,000 glycoprotein had reconstitutable Ca2+ uptake activity of 2.2 mumol/min/mg of protein. Inclusion of the Mr 55,000 glycoprotein during the reconstitution procedure did not increase the Ca2+ uptake activity of the reconstituted fraction containing the Ca2+-ATPase. This result demonstrates that the glycoprotein is not required for Ca2+ uptake.

Animals↗

PubMed1988

Virus removal or inactivation in hemoglobin solutions by ultrafiltration or detergent/solvent treatment.

Two procedures to eliminate virus infectivity from hemoglobin solutions at ambient temperature were evaluated. In the first, virus removal was assessed during the ultrafiltration of hemoglobin solutions through a membrane with a nominal molecular weight cut-off of 100,000 Daltons. The results of this study demonstrated that less than 0.1% of any virus originally spiked into the solution was detectable in the ultrafiltrate. In the second procedure the inactivation of viruses in hemoglobin solutions incubated with tri(n-butyl)phosphate mixed with sodium cholate was studied. Greater than 99% of each of the enveloped viruses tested was inactivated during the first 15 minutes of incubation with greater than 10(5) plaque forming units/ml of each being inactivated after one to six hours. No inactivation of the non-enveloped poliovirus was effected by this treatment. The data imply that both ultrafiltration and detergent/solvent incubation may reduce virus infectivity in hemoglobin solutions, but neither method yields a completely virus free product.

Cholic Acid↗

PubMedDate not supplied

Solubilisation of a novel anticonvulsant binding site from pig cortical membranes.

The present study describes the solubilisation of the novel anticonvulsant, SB-204269, binding site from pig cortical membranes. Throughout the study the binding of a close analogue of this compound, [125I]-SB-217644 (trans 6- Acetyl-4S-(3-iodobenzoylamino)-3,4-dihydro-2,2-dimethyl-2H-benzo[b ]pyran-3R-ol) was used to monitor the success of the solubilisation procedure. [125I]-SB-217644 was an ideal mechanistic tool for quantifying the binding to this novel anticonvulsant site, with a high specific activity and affinity (K(D) of 3 nmol/l). Optimum conditions for the solubilisation of this anticonvulsant binding site were investigated using a multifactorial experimental design to assess a large number of variables. Detergent type, detergent-protein ratio, absence of Mg2+ and temperature were deemed to be important factors. However, the increases observed in binding site specific activity were minimal compared with those achieved for yields. Maximum percentage yields of binding activity (25%) were achieved with a low concentration of the zwitterionic detergent, CHAPS, in the presence of a low protein concentration. This yield was further enhanced on combining mixtures of detergents. The highest recovery (37%) was achieved with a 50:50 (v:v; 1.5 x critical micelle concentration) mixture of the ionic detergent, sodium cholate, and the non-ionic detergent, MEGA-10. In summary, we report the successful solubilisation of a novel anticonvulsant binding site, identified by its selective affinity for SB-204269 and its analogues. The recovery of nearly 40% of the target binding sites from the starting material should provide a good starting point for the purification of this protein.

Animals↗