Biomedical subjects

F J Morgan

Publications and source records attributed to F J Morgan.

At least 37 records · Page 2Linked to original sources

PubMed1985

Characterization of fibrinogen New York 1. A dysfunctional fibrinogen with a deletion of B beta(9-72) corresponding exactly to exon 2 of the gene.

Fibrinogen New York 1 (NY-1) was identified in a family with a thrombotic tendency. Studies on fibrinogen NY-1 and the fibrinogen from her brother, designated NY-1a, showed that both have abnormal thrombin-nonclottable fibrinogen (50% of the total fibrinogen in NY-1 and 35-40% in NY-1a) and that the trait is heterozygous and autosomal codominant. The abnormal fibrinogen polymerizes in the presence of calcium and can be further cross-linked by Factor XIIIa. The release rates of fibrinopeptides A and B by thrombin from both (NY-1 and NY-1a) were slower than those from normal fibrinogen. Two mol of fibrinopeptide A but only 0.6-1.0 (NY-1) or 1.0-1.3 (NY-1a) mol of fibrinopeptide B were released per mol of fibrinogen. Additionally, only 1.0 (NY-1) or 1.3 (NY-1a) mol of the B beta(1-42) peptide were released by plasmin/mol. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the reduced fibrinogen revealed two protein bands in the B beta-chain region (Mr = 54,000 as compared with 57,300 for the normal). When NY-1a fibrinogen was treated with CNBr, two sizes of the NH2-terminal disulfide knot were obtained (Mr = 59,000 and 49,000). The Mr = 49,000 component is consistent with an abnormal NH2-terminal disulfide knot with two defective NH2-terminal B beta-chains. Amino acid sequence analyses demonstrated that the abnormal B beta-chain is the result of a deletion in the sequence from residues 9 to 72. This deletion corresponds exactly to exon 2 of the gene. Since this family has a thrombotic tendency, the defect in the fibrinogen may be important in the pathogenesis of thrombosis in this family.

Amino Acid Sequence↗

PubMed1985

Isolation of inhibin from bovine follicular fluid.

Bovine follicular fluid was used as a source for the isolation of gonadal inhibin, the activity of which was monitored by the dose dependent suppression of the FSH content of cultured pituitary cells. The procedures presented result in over 3000-fold purification of the starting material and the purified inhibin has an apparent molecular weight of 56000. The purified inhibin can be dissociated under reducing conditions into two subunits with molecular weights of 44000 and 14000 daltons.

Animals↗

PubMed1985

Isolation of protein S6 from rat liver ribosomes by reversed-phase high-performance liquid chromatography.

A rapid procedure for the isolation of ribosomal protein S6 from rat liver ribosomes has been developed in which proteins were separated by reversed-phase HPLC using wide-pore n-butyl-, n-octyl-, or diphenyl-bonded silica phases. Rapid processing of whole ribosomal material was achieved by the extraction of proteins in 6 M guanidinium hydrochloride and subsequent precipitation of RNA by acidification. Highly purified S6 was obtained in two chromatographic steps as shown by sodium dodecyl sulfate-gel electrophoresis, amino acid analysis, and automated microsequencing. The purification of S6 was monitored using 32P-labeled S6 as a marker which cochromatographed with unphosphorylated S6 under the low-pH elution conditions employed. Other ribosomal proteins were also purified using these reversed-phase supports, although in the case of more hydrophobic proteins such as S4 and S10 further optimization of the gradient conditions was required.

Amino Acid Sequence↗

PubMed1984

In vitro synthesis of low molecular weight proteins in human platelets: absence of labelled release products.

Isolated human blood platelets incubated at 37 degrees C in vitro incorporated labelled amino acids into compounds which included some low molecular weight (less than 80KDa) proteins, as determined by autoradiography after sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions. Non-dialysable plasma factor(s) inhibited both uptake and incorporation, which although unaffected by Actinomycin D, was inhibited partly by Chloramphenicol and almost completely by Puromycin and Cycloheximide, results which confirm that synthesis is directed by pre-existing mRNAs, some of which is mitochondrial. Assuming that the mRNA coding for proteins which are truly "platelet specific" must be present in megakaryocyte cytoplasm, we investigated the possibility that such RNA may be sufficiently stable for its translation to continue in platelets. Although leakage from platelet alpha-granules and cytoplasm during incubation was negligible and platelets retained their secretory potential we were unable to detect radiolabelled proteins in thrombin-released material after incubation. We conclude that either alpha-granule proteins are not synthesised in platelets or their megakaryocyte progenitors, or that their mRNAs become degraded by the time platelets reach the peripheral circulation. Alternatively, the mechanism which concentrates these proteins in granules does not function in circulating platelets.

Amino Acids↗

PubMed1984

Androgen receptors in breast cancer.

Androgen receptor assays have been performed on 1371 specimens of histologically confirmed primary and recurrent breast cancer. Forty-two patients who had received tamoxifen as treatment for advanced disease were assessed for objective response. Another 42 patients who had received chemotherapy were similarly studied. Patients with androgen receptor-negative tumors had a significantly poorer response rate to hormone therapy than those with receptor-positive tumors (P less than 0.05). This clinical correlation is supported by survival data of 1181 patients with primary breast cancer which showed that patients with androgen receptor-negative tumors had a highly significant trend toward shorter overall survival than those with receptor-positive tumors (P less than 0.001). Androgen receptor data added significantly to the information provided by estrogen receptor data both in terms of response to hormone treatment and survival.

Breast Neoplasms↗

PubMed1984

Sequence comparison of rat liver phenylalanine hydroxylase and its cDNA clones.

Classical phenylketonuria, an inborn error in metabolism, is caused by a deficiency of the hepatic enzyme phenylalanine hydroxylase. The identification of putative cDNA clones coding for rat liver phenylalanine hydroxylase by hybrid-selected translation has previously been reported [Robson, K. J., Chandra, T., MacGillivray, R. T. A., & Woo, S. L. C. (1982) Proc. Natl. Acad. Sci. U.S.A. 79, 4701-4705]. The authenticity of the clones, however, could not be definitively ascertained at the time because of a lack of amino acid sequence data of the enzyme in the literature. Purified rat liver phenylalanine hydroxylase was subjected to cyanogen bromide treatment, and the resulting fragments were used for N-terminal amino acid sequence analysis. The partial amino acid sequence was then compared to that deduced from an open reading frame in the nucleotide sequence of the cDNA clones. A perfect match of 17 amino acid residues was found between the two sequences following a unique methionine codon present in the nucleotide sequence, thereby providing unambiguous evidence for the identity of the rat liver phenylalanine hydroxylase cDNA clones.

Amino Acid Sequence↗

PubMed1984

High-performance liquid chromatography of amino acids, peptides and proteins. LVIII. Application of reversed-phase high-performance liquid chromatography to the separation of tyrosine-specific phosphorylated polypeptides related to human growth hormone.

Procedures for the purification of native and phosphorylated human growth hormone (hGH), S-carboxymethylated hGH, and hGH tryptic peptides, based exclusively on reversed-phase chromatography have been developed. Combinations of several volatile ion-pairing systems with small- and large-pore n-alkylsilicas were exploited in a general strategy, which allowed high recoveries of various hGH-related polypeptides from enzymatic incubations, as well as rapid desalting of samples following chemical modifications of the native protein, such as reductive alkylation in 6 M guanidine hydrochloride. The influence of the elution conditions on retention behaviour of phosphorylated hGH and its tryptic peptides in reversed-phase high-performance liquid chromatography is discussed.

Chromatography, High Pressure Liquid↗

PubMed1984

Phosphorylation of hepatic ribosomal protein S6 on 80 and 40 S ribosomes. Primary structure of S6 in the region of the major phosphorylation sites for cAMP-dependent protein kinases.

Rat liver ribosomes and 40 S ribosomal subunits were phosphorylated with the purified catalytic subunit of cAMP-dependent protein kinase. Phosphorylation of ribosomal protein S6 plateaued at around 2 mol of phosphate/mol of protein with both substrates. Peptide map analyses showed that the most prominent phosphorylation sites associated with 40 S substrates were the adjacent serines in the Arg-Leu-Ser-Ser-Leu-Arg segment of S6. The first serine residue appeared to be the preferred site as has been established previously for 80 S ribosomes (Wettenhall, R.E.H., and Cohen, P. (1982) FEBS Lett. 140, 263-269). Additional phosphorylation sites were apparent from the peptide maps. One of these was associated with the triphosphopeptide (termed T1a) having the sequence Arg-Leu-Ser-Ser-Leu-Arg-Ala-Ser-Thr-Ser-Lys. A larger fragment of S6 (termed Tlc) isolated from mild tryptic digests of whole ribosomes, consisted of the T1a sequence extended by the sequence Ser-Glu-Glu-Ser-Gln-(Lys) at the COOH terminus. A comparison of the size and chromatographic and isoelectric focusing properties of the T1a/T1c peptides and prominent tryptic peptides of S6 from insulin-stimulated hepatocytes indicated a relationship between these peptides. Thus, it appeared that some of the potential phosphorylation sites in the T1a/T1c region of S6 are phosphorylated by an insulin-regulated kinase in hepatocytes.

Amino Acid Sequence↗

PubMed1984

Purification of quadrol for microsequencing in the spinning-cup sequenator.

Treatment of Quadrol buffer with isothiocyanatophenylthiocarbamylaminoethylaminopropyl glass (DITC glass) substantially reduces the impurities observed when this buffer is used with the spinning cup sequenator. Use of DITC glass-treated Quadrol buffer permits identification of PTH-amino acids from protein degradations down to 100 pmol.

Amino Acid Sequence↗

PubMed1984

Progesterone receptors in breast cancer.

Sixty-six women who had received hormonal therapy for advanced disease were assessed for objective response to treatment. Another 51 patients who had received chemotherapy were similarly studied. Progesterone receptor was of no value as a predictor of patients unlikely to respond to hormone therapy, though it may have a role in predicting patients likely to respond favourably. The addition of progesterone receptor data to oestrogen receptor data may increase prediction of response in the ER+ range but clinicians should be cautious in their interpretation of progesterone receptor results in the ER- range. Progesterone receptor was of no value in predicting response to chemotherapy in this series. Analysis of survival data of 1731 women with primary breast cancer showed a highly significant trend toward longer survival in patients with progesterone receptor positive tumours than in those with receptor negative tumours (P less than 0.001). This trend was evident in both pre- and post-menopausal women. Even though the prognostic discrimination provided by progesterone receptor was correlated with that of oestrogen receptor, the addition of progesterone receptor data to oestrogen receptor data significantly improved prediction of survival (P less than 0.05).

Adult↗

PubMed1984

Oestradiol receptor values in breast cancer and response of metastases to therapy.

Eighty-one women who had received hormonal therapy for advanced disease were assessed for objective response to treatment. Another 73 patients who had received chemotherapy were similarly studied. Patients with ER - tumours had a significantly poorer response rate to hormonal therapy than those with ER + tumours (P less than 0.05) and a significant trend toward an increasing rate of response occurred with increasing quantitative levels of receptor (P less than 0.01). This trend was even more significant when the assay had been performed on recurrent tumour (P less than 0.001), suggesting that this provides a more accurate index of response to hormonal treatment than assays performed on the primary lesion. These results suggest that whilst the quantitative receptor level used to discriminate between 'negative' or 'positive' assays is appropriate, this division is an over-simplification. Oestrogen receptor was not helpful in predicting response to chemotherapy.

Breast Neoplasms↗

PubMed1984

The prognostic value of oestrogen receptors in breast cancer.

Survival data for 2006 women who had oestrogen receptor assay carried out on primary breast cancer tissue between 1976 and 1982 are presented. There was a significant trend to shorter survival in patients with low ER levels than in those with high ER levels (P less than 0.01). This trend was evident in both pre- and post-menopausal women. The point of maximum discrimination between prognostic groups occurred at 8 fm in premenopausal women and the four year survival rates of patients above and below this level were 84% and 48%, respectively. In post-menopausal women, maximum discrimination occurred at 90 fm, and the four year survival rates above and below this level were 82% and 64%, respectively.

Adult↗

PubMed1983

Comparison of platelet-derived growth factor prepared from release products of fresh platelets and from outdated platelet concentrates.

Platelet-derived growth factor was isolated from the release products of washed, human platelets and from freeze-thawed outdated platelet concentrates. On the basis of sodium dodecyl sulphate polyacrylamide gel electrophoresis and amino acid sequence determination we conclude that platelet derived growth factor released from platelets by the agonist thrombin (EC 3.4.4.13) is structurally similar to that isolated from lysed platelets and from platelet concentrates stored for more than 72 hr at room temperature.

Amino Acid Sequence↗

PubMed1983

Phosphorylation sites for ribosomal S6 protein kinases in mouse 3T3 fibroblasts stimulated with platelet-derived growth factor.

Platelet release products and purified platelet-derived growth factor stimulated the phosphorylation of ribosomal protein S6 in cultured mouse Balb/c 3T3 fibroblasts. The post-nuclear fraction of the stimulated cells was enriched in S6 kinase activity specific for sites resembling those phosphorylated within intact cells in response to PDGF as determined by tryptic peptide mapping. 3T3-S6 sites closely resembled those phosphorylated in S6 of rat hepatocytes stimulated with insulin and included sites for both cAMP-dependent and independent kinases.

Animals↗

PubMed1983

Hormone-receptor assays in breast cancer. A five-year experience.

Over a five-year period, from 1976 to 1981, hormone-receptor assays were performed in breast cancer specimens from 1868 women and 10 men. Oestrogen, progesterone, and androgen receptor results were distributed from zero to high levels without any clear separation into identifiable groups, which suggests that the division of results into negative and positive is an arbitrary one. The number of positive results increased significantly during the course of the study, probably due to improved tissue handling and increasing experience with assays. Over-all results showed that 61% of primary breast cancers in women compared with only 49% of secondary cancers gave positive results for oestrogen receptor. The incidence of progesterone and androgen receptors was also higher in primary cancers than in secondary cancers. In women with primary cancers, 72% of cancers were women with primary cancers, 72% of cancers were oestrogen-receptor positive in women after menopause, whereas in those before menopause only 48% were positive. Oestrogen receptor was detected in nine out of 10 male breast cancers.

Adult↗

PubMed1983

Isolation of inhibin from ovine follicular fluid using reversed-phase liquid chromatography.

This report describes the application of high performance liquid chromatography (HPLC) and associated techniques to the isolation of inhibin from ovine follicular fluid (oFF). Two modes of HPLC have been assessed. High speed gel permeation chromatography on a Waters I-125 support showed that at pH 7.0 inhibin bioactivity was associated with an approx. 80000 mol. wt protein fraction which co-eluted with the bulk of the protein in oFF and gave a yield of 30-44% of the applied activity. Reversed-phase HPLC (RP-HPLC) of a peptide-rich fraction extracted from oFF was also investigated. Using analytical liquid chromatography on octadecylsilylsilica (ODS-silica), two distinct inhibin bioactive fractions were obtained, with a resultant purification of 23-fold and 3-fold respectively and an overall recovery of 8.5%. These results demonstrate that RP-HPLC may be employed as a direct and rapid technique for the isolation of inhibin from oFF.

Animals↗

PubMed1983

Phosphorylation of human growth hormone by the epidermal growth factor-stimulated tyrosine kinase.

In the present study, we have demonstrated that human growth hormone (hGH) can be phosphorylated by the epidermal growth factor (EGF)-stimulated tyrosine kinase of A431 cell membranes. Phosphotyrosine was the predominant phosphoamino acid released from phosphorylated hGH on partial acid hydrolysis. All five tyrosine-containing tryptic peptides of hGH are also phosphorylated by the EGF-stimulated tyrosine kinase. The highest phosphate incorporation was found for peptide T4 (residues 20-38), which is distinguished by a high frequency of acidic amino acids. The phosphorylated peptides have been characterized by HPLC and two-dimensional mapping on paper. Comparison with the labeled peptides obtained on tryptic digestion of phosphorylated hGH suggests that tyrosine phosphorylation is restricted to two tryptic peptides, T4 (tyrosine-28 or -35) and T6 (tyrosine-42). It is suggested that the absence of early insulin-like activity in the naturally occurring Mr 20,000 variant of hGH, which has an internal deletion spanning residues 32-46, may be a consequence of the loss of the tyrosine phosphorylation sites at residues 35 and 42.

Amino Acid Sequence↗

PubMed1983

Isolation and complete amino acid sequence of a basic low molecular weight protein from black swan egg white.

The following complete amino acid sequence of a low molecular weight basic protein (Mr 4,454) from black swan egg white has been determined: less than Glu-Val-Arg-Lys-Tyr-Cys-Pro-Lys-Val-Gly-Tyr-Cys-Ser-Ser-Lys-Cys-Ser-Lys-Ala-Asp-Val-Trp-Ser-Leu-Ser-Ser-Asp-Cys-Lys-Phe-Tyr-Cys-Cys-Leu-Pro-Pro-Gly-Trp-Lys. There is significant homology between this protein, provisionally designated cygnin, and the NH2-terminal region of the second domain of chicken ovotransferrin. The disulfide bonds have not been assigned; however, the arrangement of half-cystines in cygnin is sufficiently different from that of the known transferrins to suggest that cygnin is derived from another gene.

Amino Acid Sequence↗