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Biomedical subjects

L Thim

Publications and source records attributed to L Thim.

162 records · Page 9Linked to original sources
PubMed1985

Primary structure of glucagon from an elasmobranchian fish. Torpedo marmorata.

Glucagon has been isolated from the pancreas of Torpedo marmorata, an elasmobranchian cartilaginous fish, and purified to homogeneity using only reverse-phase high-performance liquid chromatography. Amino acid sequence analysis indicates that the molecule differs from mammalian glucagon at position 3 (glutamic acid for glutamine), position 16 (asparagine for serine), and position 20 (lysine for glutamine). Extracts of T. marmorata intestine and brain were associated with glucagon-like immunoreactivity determined by radioimmunoassay using antisera directed against the C-terminal and N-terminal to central regions of porcine glucagon. Although elasmobranchian and teleostean fish are believed to have diverged from the main line of vertebrate evolution at about the same time, the structure of two glucagons from the teleost, Lophius americanus (anglerfish) differ from mammalian glucagon by seven and nine residues. This study supports the assertion that the structure of glucagon has been highly conserved during evolution and suggests that the considerable morphological development of the pancreas is teleosts was associated with an accelerated rate of molecular evolution.

Amino Acid Sequence↗
PubMed1985

An elasmobranchian somatostatin: primary structure and tissue distribution in Torpedo marmorata.

Extracts of brain, stomach, pancreas, and intestine from Torpedo marmorata, an elasmobranchian cartilaginous fish, contained somatostatin-like immunoreactivity. Gel filtration studies demonstrated that material with the elution volume of somatostatin-14 was the only component detected in all tissue extracts. This result contrasts with the situation in mammals where prosomatostatin is processed to multiple molecular forms in a tissue-specific manner. Somatostatin from pancreas and gut was purified to homogeneity and amino acid sequence analysis indicated that T. marmorata somatostatin from both tissues has the same structure as somatostatin-14 isolated from the higher vertebrates. Further examination of other lower vertebrate species is required in order to test the hypothesis that the ability to regulate the production of multiple forms of a regulatory peptide from a single precursor molecule developed only relatively late in evolution.

Amino Acid Sequence↗
PubMed1985

Single chain des-(B30) insulin. Intramolecular crosslinking of insulin by trypsin catalyzed transpeptidation.

Single chain des-(B30) insulin (SCI) has been synthesized from porcine insulin by trypsin in a medium with a low content of water. Trypsin catalyzes an intramolecular transpeptidation reaction in which the glycineA1 residue substitutes the alanineB30 residue, rendering a LysB29 -GlyA1 peptide link between the A- and B-chains of insulin. The insulin derivative has been purified by column chromatography and appears to be homogeneous in HPLC and disc electrophoresis. The structure was proven to be B(1-29)-A(1-21) insulin by proteolysis with Armilliaria mellea protease followed by a few steps of Edman degradation. The electrophoretic mobility indicates that SCI has a more condensed structure than that of insulin. Perfect rhombohedral crystals were obtained under conditions resembling those under which insulin crystallizes in the same form. SCI was devoid of effect in the blood sugar lowering assay in mice, the estimated potency being less than 0.1% of that of insulin.

Amino Acids↗
PubMed1984

The isolation and sequencing of human gastric inhibitory peptide (GIP).

Human GIP 1-42 and fragments of human GIP corresponding to GIP 10-42, GIP 11-42, and GIP 17-42 were isolated from acid-ethanol extracts of human small intestines with the aid of an anti-GIP serum specific for the extreme C-terminal portion of the GIP molecule. The full sequence of human GIP has been established by Edman degradation of these peptides and fragments thereof by automatic gas-phase sequencing. Human GIP differs from porcine GIP at residues 18 and 34. The sequence of human GIP is thus: (Formula: see text) Amino acid residues 18 and 34 are Arg and Ser, respectively, in porcine GIP.

Amino Acid Sequence↗
PubMed1984

Human proinsulin standards.

Two new batches of pancreatic human proinsulin have been compared with biosynthetic human proinsulin. Standards of these three proinsulin preparations were made on the basis of quantitative amino-acid analyses and compared in two proinsulin radioimmunoassays with a proinsulin standard prepared 14 years ago. The curves of the new standards were superimposable. However, they differed considerably from the curve of the old standard which proved to be only one-third of the strength of the new standards, thereby leading to a threefold over-estimation of proinsulin concentrations when the old standard is used. We conclude that the new standards should replace previously used standards.

Amino Acids↗
PubMed1984

Cyclic-AMP-dependent phosphorylation of glicentin.

Highly purified glicentin, a 69-amino-acid-residue peptide isolated from porcine intestine that contains the full sequence of glucagon and is probably biosynthetically related to glucagon, is a substrate for cyclic-AMP-dependent protein kinase in a cell-free system. Glicentin-related pancreatic peptide (residues 1-30 of glicentin) and glucagon were not phosphorylated under the same reaction conditions. It is postulated that the serine residue at position 34 of glicentin (position 2 of glucagon), that is part of the sequence Lys.Arg. His.Ser., is the probable site of phosphorylation.

Animals↗
PubMed1983

Human monocomponent insulin. Chemistry and characteristics.

The primary structure of different insulins is reviewed and the properties, identification tests, purity, potency and immunogenicity of human insulin are summerized. Novo Research Institute has developed a method, simply using an enzymatic conversion reaction to substitute the B30 alanine of porcine insulin with threonine to manufacture human insulin. This process is basically an extension of the process currently used to manufacture the Novo purified insulins which are commercially available.

Amino Acids↗
PubMed1982

Pancreatic spasmolytic polypeptide (PSP): I. Preparation and initial chemical characterization of a new polypeptide from porcine pancreas.

A novel polypeptide, named Pancreatic Spasmolytic Polypeptide (PSP), was discovered in a side-fraction from the purification of porcine insulin. PSP was prepared by two different purification methods based on combinations of precipitations, anion-exchange and cation-exchange chromatography. The highest yield obtained, 52 mg PSP/kg pancreas, indicates that the content of PSP in porcine pancreas is about half the content of insulin. Both preparations appeared to be very pure as judged by basic disc electrophoresis, isoelectric focusing, analytical gel filtration and radioimmunoassays for various polypeptides known to be present in pancreas. The PSP molecule contains 106 amino acids (MW about 11 700). PSP is an acidic (pI 4.4), non-glycosylated protein without free N-terminal amino groups, and with high contents of proline and cystine. The high content of S-S bridges (7 per molecule), an unexpected low apparent MW determined by gel filtration, and a remarkable resistance towards treatment with trypsin and chymotrypsin, point to a compact structure of the PSP molecule.

Amino Acids↗
PubMed1982

Pancreatic spasmolytic polypeptide (PSP): II. Radioimmunological determination of PSP in porcine tissues, plasma and pancreatic juice.

A radioimmunoassay (RIA) for the determination of porcine Pancreatic Spasmolytic Polypeptide (PSP) has been developed. The antisera raised in rabbits were sensitive to 5 pgequiv. of PSP in a volume of 100 microliter. Immunoreactive PSP (IR-PSP) has been determined in extracts from 22 porcine organs. Pancreas was found to be the only organ containing substantial amounts of IR-PSP (0.1 mgequiv. IR-PSP/g wet weight). The fasting porcine plasma level of IR-PSP was about 10 ngequiv./ml, corresponding to 850 pM. The concentration of IR-PSP in porcine pancreatic juice varied from 0.2 mugequiv./ml in the fasting state to 46-116 mugequiv./ml after stimulation with pancreozymin or secretin. A linear correlation was found between the exocrine secretion of IR-PSP and total protein.

Animals↗
PubMed1982

Pancreatic spasmolytic polypeptide (PSP): III. Pharmacology of a new porcine pancreatic polypeptide with spasmolytic and gastric acid secretion inhibitory effects.

Pancreatic spasmolytic Polypeptide (PSP) is a new porcine pancreatic polypeptide, which inhibits gastrointestinal motility and gastric acid secretion in laboratory animals after parenteral as well as oral administration. (1) PSP inhibits the amplitude of electrically stimulated contractions of the isolated guinea pig ileum. PSP's inhibitory effect is antagonized by phentolamine, but not by yohimbine. (2) PSP inhibits the motility of isolated guinea pig intestinal segments after intraluminal dosing. (3) PSP reduces intestinal motility in rabbits in vivo after intravenous and intraluminal administration, and in mice in vivo after subcutaneous injection. (4) PSP delays absorption of protein hydrolysate when it is administered orally in capsules to pigs and to pancreatectomized dogs. (5) PSP inhibits pentagastrin induced gastric acid secretion in rats after oral administration and in cats after subcutaneous and oral administration. The mechanism of action of PSP has so far not been finally elucidated. It seems likely that PSP interferes with endogenous acetylcholine release. Furthermore it might act by release of somatostatin from somatostatin cells in the gastrointestinal tract. It may have a direct or an indirect stimulant effect on alpha 2-receptors.

Administration, Oral↗
PubMed1981

Relationship of glicentin to proglucagon and glucagon in the porcine pancreas.

We have previously isolated from porcine small intestine a peptide known as glicentin. The C-terminal portion of glicentin consists of the sequence of glucagon extended at its C terminus by an octapeptide, and differs slightly from the sequence of a proposed fragment of proglucagon. Glicentin-like material has been demonstrated in the pancreatic A cell, wherein it is located in the periphery of the secretory granules, whereas glucagon is located in the centre of the granules. To study the relationship of glicentin to the biosynthesis of glucagon, we have now investigated the glucagon-like and glicentin-like peptides in extracts and perfusates of the porcine pancreas. Our findings that a peptide with glicentin-like immunoreactivity, and intermediate in size between glicentin and glucagon, is secreted synchronously with glucagon suggest that this glicentin-related peptide is a major cleavage product of proglucagon.

Animals↗
PubMed1981

The primary structure of porcine glicentin (proglucagon).

The primary structure of porcine glicentin has been established. The molecule consists of 69 amino acid residues and has a molecular weight of 8128. The sequence of glicentin 1-30 represents the glicentin-related pancreatic peptide (GRPP) previously isolated from porcine pancreas. The sequence 33-61 represents the full sequence of glucagon and the sequence 64-69 is a C-terminal hexapeptide. These three sequences, GRPP, glucagon and the hexapeptide are linked by two Lys-Arg pairs which probably represent the sites for post-synthetic enzymatic cleavages. Glicentin thus fulfils the structural requirements for being proglucagon.

Amino Acid Sequence↗
PubMed1981

The amino acid sequence of porcine glicentin.

Glicentin or gut GLI-1 has previously been isolated from porcine small intestine. On the basis of the available chemical data, the molecule was thought to contain 100 amino acid residues. A redetermination of the amino acid composition of the molecule has shown it to contain 69 amino acid residues, and the full sequence has been established. The sequence of glicentin can be outlined as: GRPP1--30-Lys-Arg-Glucagon33--61-Lys-Arg-Hexapeptide64--69 where GRPP1--30 probably corresponds to the glicentin related pancreatic peptide previously isolated from porcine pancreas. In the pancreas, the two dibasic sequences (Lys31-Arg32 and Lys62-Arg63) probably represent sites of post-synthetic enzymatic cleavages by analogy with the two dibasic sequences of proinsulin. Glicentin thus fulfills the structural requirements for being all or a part of porcine proglucagon. In the intestine, glicentin could be the precursor of oxyntomodulin, a small molecular weight gut GLI presumably identical to glicentin 33--69, i.e., glucagon extended at the C-terminal end by an octapeptide.

Amino Acid Sequence↗
PubMed1978

Heparin-sepharose affinity chromatography of human post-heparin plasma. Characterization of the elution pattern with immunoelectrophoretic methods.

Immunochemical methods including fused rocket-, crossed-, and tandem-crossed immunoelectrophoresis, have been used to characterize the elution pattern from heparin-Sepharose affinity, chromatography of human post-heparin plasma. Hepatic triglyceride lipase (H-TGL) but not lipoprotein lipase (LPL) could be visualized with beta-naphthyl acetate after immunoelectrophoresis. Two proteins were found to elute together with the lipolytic enzymes. The amino acid composition of fractions containing these proteins was nearly identical to that of antithrombin III. These results indicate that the removal of antithrombin III is the major problem in the purification of H-TGL and LPL from human post-heparin plasma by heparin-Sepharose affinity chromatography.

Chromatography, Affinity↗
PubMedDate not supplied

Neuropeptide K-(1-24)-peptide: storage and release by carcinoid tumors.

An antiserum directed against the COOH-terminal region of neuropeptide K-(1-24)-peptide that shows only 0.5% reactivity with neuropeptide K has been used in radioimmunoassay to study the posttranslation processing of human beta-preprotachykinin. A primary midgut carcinoid tumor contained high concentration of substance P (2970 pmol/g), neurokinin A (3660 pmol/g) and neuropeptide K-(1-24)-peptide (3430 pmol/g) but only a very low concentration (less than 5 pmol/g) of intact neuropeptide K. Neuropeptide K-(1-24)-peptide was also detected in extracts of metastatic tumor tissue from four patients with midgut carcinoid tumors. The amino acid sequence of tumor neuropeptide K-(1-24)-peptide was identical to that predicted from the nucleotide sequence of a human beta-preprotachykinin cDNA. The fasting plasma concentration of neuropeptide K-(1-24)-peptide was elevated in a patient with the carcinoid syndrome (821 fmol/ml compared with less than 18 fmol/ml in healthy subjects) and rose approximately 2-fold after intravenous pentagastrin. The study has demonstrated that the Lys25-Arg26 bond in neuropeptide K (corresponding to Lys96-Arg97 in the precursor) is an important processing site in human beta-preprotachykinin.

Aged↗
PubMedDate not supplied

Neuropeptide Y-related peptides from the pancreas of a teleostean (eel), holostean (bowfin) and elasmobranch (skate) fish.

Homologous peptides belonging to the pancreatic polypeptide (PP) family were isolated from the pancreas of a teleostean fish, the American eel (Anguilla rostrata), an holostean fish, the bowfin (Amia calva) and an elasmobranch fish, the skate (Raja rhina), and their primary structures were determined. The peptides show stronger homology to neuropeptide Y, particularly in their COOH-terminal regions, than to peptide YY or pancreatic polypeptide and contain an alpha-amidated COOH-terminal tyrosine residue. The skate peptide Tyr-Pro-Pro-Lys-Pro-Glu-Asn-Pro-Gly-Asp10-Asp-Ala-Ala-Pro-Glu-Glu- Leu-Ala-Lys- Tyr20-Tyr-Ser-Ala-Leu-Arg-His-Tyr-Ile-Asn-Leu30-Ile-Thr-Arg- Gln-Arg-Tyr-NH2 represents the first member of the PP family to be isolated from a cartilaginous fish. The primary structure of the pancreatic PP family peptide has been more strongly conserved among the phylogenetically more ancient holostean and elasmobranch fishes than among the teleosts. A comparison of the primary structures of all PP family peptides supports the hypothesis and evolution has acted to conserve features of tertiiary structure in the molecules (e.g., the polyproline- and alpha-helices) rather than individual amino acid residues.

Amino Acid Sequence↗
PubMedDate not supplied

Rainbow trout (Oncorhynchus mykiss) neuropeptide Y.

Neuropeptide Y (NPY) has been isolated from brain extracts of the rainbow trout (Oncorhynchus mykiss) and subjected to structural analyses. Plasma desorption mass spectroscopy estimated the molecular mass of the purified peptide as 4303.9 Da. Automated Edman degradation unequivocally established the sequence of a 36 amino acid residue peptide as: Tyr-Pro-Pro-Lys-Pro-Glu-Asn-Pro-Gly-Glu-Asp-Ala-Pro-Pro-Glu-Glu-Leu-Ala- Lys- Tyr-Tyr-Thr-Ala-Leu-Arg-His-Tyr-Ile-Asn-Leu-Ile-Thr-Arg-Gln-Arg-Tyr. The molecular mass calculated from this sequence (4304 Da) is consistent with that obtained by mass spectroscopy. The presence of a C-terminal amide was established by radioimmunoassay. Rainbow trout NPY is identical in primary structure to coho salmon (Oncorhynchus kisutch) pancreatic polypeptide (PP). These data may indicate that, in this group of salmonid fishes, a single member of the NPY/PP peptide family is expressed in both neurons and peripheral endocrine cells.

Amino Acid Sequence↗