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Biomedical subjects

S Birken

Publications and source records attributed to S Birken.

At least 73 records · Page 4Linked to original sources
PubMed1984

A human myeloma immunoglobulin G binding four moles of calcium associated with asymptomatic hypercalcemia.

A calcium-binding immunoglobulin G (IgG1 lambda RUP) was identified in the serum of a patient with multiple myeloma, asymptomatic hypercalcemia, and a normal ionized serum calcium. Calcium binding by IgGRUP was confirmed by two-dimensional electrophoresis with calcium-45 and equilibrium dialysis. Amino acid analyses indicated an unusually high number of glutamic (or glutamine) residues in the L chain and Fab fragment but no detectable gamma-carboxyglutamic acid. As determined by equilibrium dialysis with 45Ca, the intact IgGRUP and its Fab fragments bound calcium at an optimum pH of 7.4. There was minimal binding of calcium to H chains and no binding by L chains or the Fc fragment. Recombination of H and L chains partially restored the binding activity. By Scatchard analysis, the binding affinity (Kd) of IgGRUP was 1.7 X 10(-3) M and the binding capacity was 4 mol of calcium/mol of IgG. The binding of 4 mol of calcium/mol of IgG is twice that reported previously for two other calcium-binding myeloma proteins and suggests unique properties of IgGRUP.

Aged↗
PubMed1984

Family studies of the genetic abnormality in transthyretin (prealbumin) in Portuguese patients with familial amyloidotic polyneuropathy.

Amyloid deposits in several heredofamilial forms of amyloidosis are chemically related to transthyretin (TTR, the protein usually referred to as prealbumin). A genetically abnormal TTR may be involved. Studies were conducted on TTR isolated from sera of patients with familial amyloidotic polyneuropathy (FAP), and on amyloid fibril protein (AFp) isolated from tissues of two Portuguese patients who died with FAP. AFp, purified by affinity chromatography on retinol-binding protein (RBP), resembled plasma TTR in forming a stable tetrameric structure, and in its binding affinities for both thyroxine and RBP. Purified AFp was found to comprise a TTR variant with a methionine for valine substitution at position 30. This conclusion was based upon studies that included: (i) comparative peptide mapping by reverse-phase high-performance liquid chromatography after trypsin digestion; (ii) cyanogen bromide (CNBr) cleavage studies; and (iii) amino acid microsequence analysis of selected tryptic and CNBr peptides. The variant TTR was also found to be present in serum samples from FAP patients, along with larger amounts of normal TTR. An effective, small-scale procedure was developed to determine whether or not the variant TTR was present in the plasma of an individual subject. This procedure involved isolation of TTR by affinity chromatography on RBP, followed by CNBr cleavage, and analysis for the presence of specific aberrant CNBr peptides. Studies with six kindreds, including 21 asymptomatic children of 6 patients with FAP, showed that the "abnormal" TTR can be detected and used as a preclinical marker of the disease in affected children of patients with FAP. It is likely that the variant TTR represents a point mutation within the TTR structural gene, and that the normal and mutant genes act as co-dominant alleles at a single locus in FAP. The distribution of the mutant TTR within the six families was consistent with the autosomal dominant mode of inheritance of FAP. The mutant TTR apparently selectively deposits in tissues as the amyloid characteristic of the disease.

Adolescent↗
PubMed1984

Amyloid fibril protein in familial amyloidotic polyneuropathy, Portuguese type. Definition of molecular abnormality in transthyretin (prealbumin).

Amyloid fibril protein in patients with familial amyloidotic polyneuropathy is known to be chemically related to transthyretin (TTR), the plasma protein that is usually referred to as prealbumin. A genetically abnormal TTR may be involved in this disease. Studies were conducted on amyloid fibril protein (AFp) isolated from tissues of two Portuguese patients who died with familial amyloidosis, and on TTR isolated from sera of patients with this disease. AFp, purified by affinity chromatography on retinol-binding protein linked to Sepharose, resembled plasma TTR in forming a stable tetrameric structure, and in its binding affinities for both thyroxine and retinol-binding protein. The structural studies included: (a) comparative peptide mappings by reverse-phase high performance liquid chromatography (HPLC) after trypsin digestion; (b) cyanogen bromide cleavage studies; and (c) amino acid microsequence analysis of selected tryptic and CNBr peptides. On the basis of the known amino acid sequence of TTR, comparative tryptic peptide maps showed the presence of a single aberrant tryptic peptide (peptide 4, residues 22-34) in AFp as compared with TTR. This aberrant peptide contained a methionine residue, not present in normal tryptic peptide 4. CNBr cleavage of AFp produced two extra peptide fragments, which were demonstrated, respectively, by HPLC analysis and by sodium dodecyl sulfate-gel electrophoresis. Sequence analyses indicated the presence of a methionine-for-valine substitution at position 30 in AFp as compared with TTR. Thus, the purified amyloid fibril protein comprised a TTR variant with a methionine-forvaline substitution at position 30. A single nucleotide change in a possible codon for valine 30 could explain the substitution. The variant TTR was also present in the TTR isolated from the pooled sera of amyloidoses patients, together with larger (four- to six-fold) amounts of the normal TTR. Thus, in these patients, the variant TTR was circulating in plasma, along with larger amounts of normal TTR. We suggest that the variant TTR represents the specific biochemical cause of the disease, and that this abnormal form of TTR selectively deposits in tissues as the amyloid characteristic of the disease.

Amino Acid Sequence↗
PubMed1984

Use of a highly sensitive and specific immunoradiometric assay for detection of human chorionic gonadotropin in urine of normal, nonpregnant, and pregnant individuals.

A highly sensitive and specific two-site immunoradiometric assay (IRMA) for hCG has been developed and applied to the detection of the hormone in the urine of normal nonpregnant and pregnant individuals. The IRMA uses a solid phase coupled monoclonal antibody to the hCG beta-subunit for extraction of hormone from urine. The hCG extracted is then directly quantified by the binding of an affinity purified and radiolabeled rabbit antibody that reacts with the unique COOH-terminal peptide region of the hCG beta-subunit. The assay is capable of reliably and accurately measuring as little as 0.01 ng hCG/ml urine without interference from hLH. Assays of urine from normal men and nonpregnant women of reproductive age indicated that most individuals did not have detectable levels of hCG immunoreactivity, although a minority had minute amounts, with a mean value of approximately 0.01 ng hCG/mg creatinine. In contrast, all normal menopausal women studied had easily detectable levels of hCG immunoreactivity in their urine, with a mean value of 0.123 ng hCG/mg creatinine. A study of the excretion of hCG from three men injected with hormone for treatment of infertility indicated that after the first 24 h, hCG was cleared with a single exponential rate and was detectable to a level of 0.01 ng/ml. Application of the IRMA to measurements of hCG in the urine of two artificially inseminated patients indicated that the method was capable of detecting pregnancy as early as 9 days postovulation. The extreme sensitivity and specificity of the IRMA for urinary hCG in conjunction with the simplicity of assay performance and specimen collection should provide a substantial advantage over currently available methods for detection of early pregnancy and tumor monitoring.

Adult↗
PubMed1984

Effect of desialylation of human chorionic gonadotropin on its metabolic clearance rate in humans.

We determined the MCR and other kinetic parameters of desialylated hCG (as-hCG) in eight subjects (four men and four women), aged 20-47, using a single injection of 1 mg of a highly purified preparation. The plasma disappearance curve was fitted to a single exponential equation, with a half-time of disappearance of 3.56 +/- 0.62 min (mean +/- SD) and an initial volume of distribution of 1790 +/- 582 ml/m2. The mean MCR of as-hCG was 349 +/- 96 ml/min X m2. The mean transit time of as-hCG was 373 times shorter than that of hCG. The rapid plasma disappearance of as-hCG could not be accounted for by renal clearance into urine, since less than 0.05% of the as-hCG injected was excreted in the urine, nor could it be explained by degradation by plasma proteases, since as-hCG was not appreciably degraded during incubation in human serum at 37 C for 3 h. These data show that desialylation markedly accelerates the MCR of hCG in the human in spite of reduced renal excretion and, therefore, indicate the presence of a highly active pathway for uptake of desialylated glycoproteins.

Adult↗
PubMed1984

Chemistry of human choriogonadotropin.

Human choriogonadotropin is the placental member of the homologous glycoprotein hormone family which includes the pituitary hormones lutropin, follitropin and thyrotropin all of which share a common alpha subunit but different target-specific beta subunits. HCG differs chemically from the pituitary hormones by its higher carbohydrate content, high sialic acid content, lack of sulfation, longer plasma half-Life and capability to survive passage through the kidney essentially intact. Since crystallographic models are not available, hormone topography has been explored by immunochemical methods using both polyclonal and monoclonal antibodies. Studies of the immunochemistry of the beta COOH-terminal region have shown that determinants dependant upon carbohydrate content exist within this region. Such work also led to development of extremely sensitive and specific immunoassays for measurement of the hormone.

Amino Acid Sequence↗
PubMed1984

Unusual molecular forms of hCG in gestational trophoblastic neoplasia.

We have observed unusual forms of human choriogonadotropin (hCG) in the urine of patients with gestational trophoblastic neoplasia. These molecules were of two types: Carboxyterminal fragments of hCG beta-subunit and desialylated forms of hCG. Substantial amounts of these materials were found in the urines of 83% and 50%, respectively, of the patients (n = 12) studied. In contrast, negligible quantities were found in the urine of 14 healthy pregnant women. Also, these unusual forms were not detectable in normal controls given an infusion of hCG. These findings indicate that these forms have clinical potential for diagnosing gestational trophoblastic neoplasia and differentiating it from normal pregnancy.

Chemical Phenomena↗
PubMed1983

Monoclonal antibody to the region of fibronectin involved in cross-linking to human fibrin.

A cross-link-containing fragment (alpha XLCNBr), derived from the alpha-polymer component of human fibrin following CNBr digestion, has been isolated and characterized. NH2-terminal sequence studies of three alpha XLCNBr derivatives, each prepared by a different method, indicate that the A alpha-chain regions comprised of residues 241-476 (CNBr VIII) and 518-584 (CNBr X) are the major constituents of the cross-linked fragments examined. Evidence for at least two additional sequences suggests that A alpha 208-235 (CNBr V) and A alpha 585-610 (CNBr XI) may have auxiliary roles in alpha-polymer formation. When alpha XLCNBr was used as an immunogen for the production of murine hybridoma cell lines, two groups of antibodies were obtained. The majority of supernatants from primary hybridoma cultures did not discriminate between fibrinogen and alpha XLCNBr and appeared to contain antibodies directed against determinants within either CNBr VIII or CNBr X [see Ehrlich, P. H., Sobel, J. H., Moustafa, Z. A., & Canfield, R. E. (1983) Biochemistry (following paper in this issue)]. Several supernatants from primary hybridoma cultures, however, did exhibit significant binding toward the alpha-polymer derivative in the absence of demonstrable immunoreactivity toward either highly purified fibrinogen or its A alpha-chain peptides, CNBr VIII and CNBr X.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1983

Hemoglobin-binding site on haptoglobin probed by selective proteolysis.

Selective proteolysis has been used to delineate the hemoglobin-binding site on haptoglobin heavy chain. Haptoglobin was cleaved specifically by plasmin, trypsin, chymotrypsin, staphylococcal protease, and thermolysin. Haptoglobin-hemoglobin complex was treated with these enzymes to determine which sites were protected from cleavage by the hemoglobin. The modified haptoglobins were tested for changes in their hemoglobin and hemoglobin alpha chain-binding properties. The sites of proteolytic cleavage were identified from the newly generated NH2 termini by automated Edman degradation amino acid-sequencing techniques. The results suggest that residues 128 through 131, 136 and 137, as well as 9 and 10 of the heavy chain may be involved in the binding of hemoglobin. On the other hand, residues 159 and 160, which lie in the 17-residue additional loop that is unique to haptoglobin among its homologous serine protease family, and residues 73 and 74, which lie close to the carbohydrate-binding residues, appear to be remote from the hemoglobin-binding site.

Amino Acid Sequence↗
PubMed1983

Preferential masking by the receptor of immunoreactive sites on the alpha subunit of human choriogonadotropin.

125I-Labeled human choriogonadotropin (125I-hCG) bound to rat ovarian receptor was solubilized in Triton X-100. By using increasing concentrations of nine different antisera specific for the individual subunits of human choriogonadotropin (hCG), free 125I-hCG or 125I-hCG-receptor complex was precipitated by double-antibody technique. The ability of any antiserum to bind to the hormone-specific beta subunit was not affected by hCG binding to receptor, suggesting that this subunit is not directly involved with the receptor in the final state of the hormone-receptor complex. In contrast, every antiserum specific for the alpha subunit was dramatically inhibited in binding to the solubilized 125I-hCG-receptor complex. These results suggest that the alpha subunit directly interacts with the receptor, thereby masking immunoreactive sites normally available on the free hormone. Because a number of reports describe binding activity of high concentrations of immunopurified beta subunits of hCG, we propose a two-step model for the binding of hCG to receptor and postulate separate and distinct roles for the subunits. We propose that the binding of hCG to the receptor involves a specific low-affinity initial interaction of the beta subunit with the receptor that activates a second site for the high-affinity binding of alpha subunit and stabilization of the hormone-receptor complex.

Animals↗
PubMed1983

Characterization of a carboxyterminal peptide fragment of the human choriogonadotropin beta-subunit excreted in the urine of a woman with choriocarcinoma.

We have observed low-molecular weight carboxyterminal fragments of the human choriogonadotropin (hCG) beta-subunit in the urines of several women with choriocarcinoma, and we have characterized one fragment in detail. Its apparent molecular weight by gel chromatography on Sephadex G-100 was 14,200. The fragment was not adsorbed to concanavalin A-Sepharose, indicating that it lacked the asparagine-linked carbohydrate groups of intact hCG beta. It was active in radioimmunoassays (RIA) using antisera either to the hCG beta carboxyterminal peptide (CTP) or to the desialylated hCG beta CTP (hCG beta as-CTP), indicating the presence of not only the hCG beta carboxyterminus but also desialylated O-serine-linked carbohydrate side chains on the fragment. It lacked luteinizing hormone/choriogonadotropin radioreceptor activity and hCG beta conformational immunoreactivity (SB6 RIA). On Sephadex G-100 gel chromatography, the elution profiles of this fragment and the hCG beta as-CTP(115-145) prepared by trypsin digestion of as-hCG were essentially indistinguishable (apparent molecular weights 14,200 and 14,000, respectively). The immunological characteristics of the fragment in both hCG beta CTP and hCG beta as-CTP RIA were indistinguishable from those of the hCG beta as-CTP(115-145) glycopeptide. Carboxyterminal fragments of hCG beta were evident in urine specimens obtained from 10 of 11 patients with choriocarcinoma but not in those obtained from normal subjects who were given an intravenous infusion of highly purified hCG. Of six pregnant women, only the one at term excreted carboxyterminal fragments of hCG beta and then only in trace amounts. We conclude that hCG beta carboxyterminal fragments, including one that is indistinguishable from the tryptic glycopeptide hCG beta as-CTP(115-145), can occur naturally in the urine of patients with choriocarcinoma.

Choriocarcinoma↗
PubMed1983

Presence of an abnormal transthyretin (prealbumin) in Portuguese patients with familial amyloidotic polyneuropathy.

In summary, these studies have demonstrated the following: 1) plasma TTR levels are significantly decreased in Portuguese patients with FAP; 2) FAP-TTR is indistinguishable from normal TTR with regard to a wide range of physiochemical properties; 3) RBP levels and vitamin A transport appear to be normal in FAP patients; 4) an abnormal TTR with a methionine for valine substitution at position 30 is found as amyloid protein in tissues of patients with FAP; and 5) this abnormal TTR is present in small amounts in plasma of patients with FAP. Based on these observations, we postulate that this abnormal form of TTR selectively deposits in tissues as the amyloid characteristic of the disease.

Adult↗
PubMed1982

Absence of the COOH-terminal peptide on ectopic human chorionic gonadotropin beta-subunit (hCG beta).

A comparison of the physical properties and immunoreactivity of ectopic hCG beta from cultured cervical cancer cells and standard hCG beta was performed. The heterogeneity of the preparations was examined by ion-exchange chromatography, and the molecular size by analytical gel filtration. Two distinct forms of ectopic hCG beta were identified. One form was indistinguishable from standard hCG beta, while the other, although larger, lacked the characteristic COOH-terminal peptide (CTP). This was shown by the failure of antisera specific for determinants on the cTP to recognize this molecule, and by the apparent absence of the 0-linked oligosaccharides and thermolysin cleavage site normally found in this region.

Agglutinins↗
PubMed1981

Improved monitoring of gestational trophoblastic neoplasia using a highly sensitive assay for urinary human chorionic gonadotropin.

A radioimmunoassay for urinary human chorionic gonadotropin (hCG) which used antiserum to the hCG beta carboxy-terminal peptide was employed to detect hCG production in women with gestational trophoblastic disease. The urinary hCG assay was considerably more effective in detecting increased hCG production, and, hence, persistent tumor burden, than the commonly used serum hCG beta subunit radioimmunoassay. In two women treated for choriocarcinoma, urinary hCG levels remained distinctly elevated for 2 to 6 weeks, even though the serum hCG level was undetectable; in both cases, the urinary hCG radioimmunoassay reliably predicted the eventual reappearance of hCG in the serum. In six women whose disease remitted, the urinary hCG level declined to the normal range during chemotherapy, and remained there afterward. In eight patients who had been in remission for periods of from 1 to 14 years, urinary hCG levels were in the normal range. We conclude that this highly sensitive and reliable method for the detection of hCG production improves the ability to diagnose persistent trophoblastic disease.

Adult↗
PubMed1981

The amino acid sequences of the prepeptides contained in the alpha and beta subunits of human choriogonadotropin.

The amino acid sequences of the NH2-terminal extension prepeptides of the alpha and of the beta subunits of human choriogonadotropin have been determined by translation of first trimester placental mRNA in the presence of individually labeled amino acids. The structure of the prepeptide of the alpha subunit is: -24 -23 -22 -21 -20 -19 -18 -17 -16 -15 Met- Asp- Tyr- Tyr- Arg- Lys- Tyr- Ala- Ala- Ile -14 -13 -12 -11 -10 -9 -8 -7 -6 -5 Phe- Leu- Val- Thr- Leu- Ser- Val- Phe- Leu- His -4 -3 -2 -1 Val- Leu- His- Ser The structure of the prepeptide of the beta subunit is: -20 -19 -18 -17 -16 -15 -14 -13 -12 -11 Met- Glu- Met- Phe- Gln- Gly- Leu- Leu- Leu- Leu -10 -9 -8 -7 -6 -5 -4 -3 -2 -1 Leu- Leu- Leu- Ser- Met- Gly- Gly- Thr- XXX- Ala In the presence of membranes, the beta prepeptide is cleaved to a protein containing the authentic NH2-terminal sequence of the secreted form of the beta subunit. Cleavage of pre-alpha subunit also yields a product containing the NH2-terminal region of the secreted form.

Amino Acid Sequence↗