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Biomedical subjects

D Hawke

Publications and source records attributed to D Hawke.

At least 19 recordsLinked to original sources
PubMed1991

Immunoglobulin variable-region-like domains of diverse sequence within the major histocompatibility complex of the chicken.

The highly polymorphic B-G antigens are considered to be part of the major histocompatibility complex (MHC) of the chicken, the B system of histocompatibility, because they are encoded in a family of genes tightly linked with the genes encoding MHC class I and class II antigens. To better understand these unusual MHC antigens, full-length B-G cDNA clones were isolated from B21 embryonic erythroid cell cDNA library, restriction-mapped, and sequenced. Five transcript types were identified. Analysis of the deduced amino acid sequences suggests that the B-G polypeptides are composed of single extracellular domains that resemble immunoglobulin domains of the variable-region (V) type, single membrane-spanning domains typical of integral membrane proteins, and long cytoplasmic tails. Sequence diversity among the five transcript types was found in all domains, notably including the B-G immunoglobulin V-like domains. The cytoplasmic tails of the B-G antigens are made up entirely of units of seven amino acid residues (heptads) that are typical of an alpha-helical coiled-coil conformation. The heptads vary in number and sequence between the different transcripts. The presence within B-G polypeptides of polymorphic immunoglobulin V-like domains warrants further investigations to determine the degree and nature of variability within this domain in these unusual MHC antigens.

Amino Acid Sequence↗
PubMed1985

Primary structure of a glycosylated DNA-binding domain in human plasma fibronectin.

The complete amino acid sequence of a DNA-binding domain isolated from human plasma fibronectin after limited trypsin digestion has been obtained. It contains 132 amino acids and one biantennary glycosyl unit at residue 104, for an estimated Mr of 16,931. The fragment can be purified by a two-step procedure consisting of DNA-affinity chromatography and reverse-phase high performance liquid chromatography. It can also be purified by heparin-affinity chromatography. The domain is unusual in its susceptibility to tryptic-like cleavages even by neutral or aromatic residue-specific proteases. It has no cysteine residues and is predicted to favor a beta-sheet structure by Chou and Fasman analysis. Based on this analysis we have proposed a model which exhibits a clustering of aromatic and basic residues, consistent with similar involvement of basic and aromatic residues in other DNA-binding proteins. The net charge of the domain at neutral pH (+1, without sialic acid) argues against a nonspecific charge interaction with polyanionic macromolecules such as DNA and heparin. Internal sequence repeats occur at intervals of 30, 60, and 90 residues, thus suggesting a maximum size for a repetitive building block which gave rise to this domain.

Amino Acid Sequence↗
PubMed1985

Purification of somatostatin from frog brain: coisolation with retinal somatostatin-like immunoreactivity.

Somatostatin-like immunoreactivity (SLI) was purified from frog brain and retina, and the structure of the brain peptide was determined. Frog brain (101 g) and retinal (45 g) tissues were extracted with 3% acetic acid, yielding 9.6 and 0.44 nmol of SLI, respectively. SLI was further purified by chromatography on a somatostatin immunoaffinity column followed by sequential application to reverse-phase C-18 HPLC columns. The brain and retinal peptides, purified roughly 100,000-fold with net yields of 7.5 and 2.3%, respectively, appeared identical in the final steps of purification. The amino acid sequence of brain SLI, as determined by a gas-phase automated Edman degradation technique, was as follows: Ala-Gly-(Cys)-Lys-Asn-Phe-Phe-Trp-Lys-Thr-Phe-Thr-Ser-(Cys). Our data indicate that despite structural variations in somatostatins of other lower vertebrates, the amino acid sequence of frog brain and, by deduction, retinal SLI is identical to that of somatostatin tetradecapeptide. These findings support the physiological relevance of studies directed at elucidating the neurotransmitter function of somatostatin using the well-established models of frog brain and retina.

Amino Acid Sequence↗
PubMed1985

A nonamidated peptide homologous to porcine peptide YY and neuropeptide YY.

A 37-residue peptide has been purified from the endocrine pancreas of the anglerfish. The first 36 residues were sequenced by gas phase Edman degradation. The sequence at the carboxyl-terminus was determined by sequencing the carboxyl-terminal tryptic dipeptide. The sequence of the peptide, which is consistent with the amino acid composition, was determined to be: Y X P X P X K X P X E X T X P X G X S X N X A X S X P X E X D X W X A X S X Y X Q X A X A X V X R X H X Y X V X N X L X I X T X R X Q X R X Y X G. Fast atom bombardment/mass spectrometry of the peptide identified a molecular ion with an average mass of 4221.3, in good agreement with the theoretical mass based upon the determined amino acid sequence. The peptide has an equal degree of sequence identity to both porcine neuropeptide YY (64%) and gastrointestinal peptide YY (64%), but less sequence identity to porcine pancreatic polypeptide (47%). Unlike the related mammalian peptides, the major form of the anglerfish peptide terminates in tyrosyl-glycine rather than tyrosineamide.

Amino Acid Sequence↗
PubMed1984

Anglerfish preprosomatostatin II is processed to somatostatin-28 and contains hydroxylysine at residue 23.

A peptide fraction containing two 28-residue somatostatins, both products of the anglerfish somatostatin II gene, has been isolated, characterized, and subjected to amino acid sequence analysis. The structural data indicate that one of the two forms of the 28-residue peptide contains 5-hydroxylysine. Hydroxylysine was identified in an acid hydrolysate of somatostatin-28 by gas chromatography/mass spectrometry. Fast-atom bombardment mass spectrometry indicated that the two forms of somatostatin-28 have molecular weights of 3220 and 3204, representing the hydroxylated and nonhydroxylated peptides, respectively. The location of the hydroxylated lysine was deduced by analysis of proteolytic fragments to be position 23. This represents the first observation of a hydroxylated peptide hormone and one of the few reported occurrences of hydroxylysine in non-collagen proteins.

Amino Acid Sequence↗
PubMed1984

Amino terminal fragments of human progastrin from gastrinoma.

Two peptides which copurified from a human gastrinoma were found to correspond to the amino acid sequence deduced for the amino terminal portion of human and porcine progastrin. The sequence of peptide A is Ser-Trp-Lys-Pro-Arg-Ser-Gln-Gln-Pro-Asp-Ala-Pro-Leu-Gly-Thr-Gly-Ala-Asn- Arg-Asp-Leu-Glu-Leu which is identical to an amino terminal portion of human progastrin. The sequence of peptide. B is identical to that of peptide A except it is missing the first five amino acids. If peptide A corresponds to the amino terminus of progastrin, the signal peptidase cleaves at an Ala-Ser bond.

Amino Acid Sequence↗
PubMed1984

The amino acid sequence of a fluorescein-labeled peptide from the active site of (Na,K)-ATPase.

(Na,K)-ATPase in an active-transport protein that couples the energy obtained from the hydrolysis of ATP to the transport of Na+ and K+ across animal cell membranes. In order to investigate the enzymatic mechanism of this activity, a peptide derived from the ATP-binding site of (Na,K)-ATPase has been purified and its amino acid sequence has been determined. The peptide was identified by the covalent incorporation of a fluorescent probe, fluorescein 5'-isothiocyanate, into the active site before trypsin digestion of the protein. The labeling of (Na,K)-ATPase by fluorescein 5'-isothiocyanate was associated with the irreversible inhibition of enzymatic activity, and both the labeling of the tryptic peptide and inhibition of activity were prevented when the reaction was performed in the presence of ATP. An apparent KD of 5.7 microM was calculated when the reaction between (Na,K)-ATPase and fluorescein 5'-isothiocyanate was performed under pseudo first-order conditions. The amino acid sequence of the active-site peptide, His-Leu-Leu-Val-Met-Lys-Gly-Ala-Pro-Glu-Arg, is similar to the sequence of a fluorescein-labeled peptide derived from the active site of the sarcoplasmic reticulum Ca2+-transport ATPase (Mitchinson, C., Wilderspin, A. F., Trinaman, B. J., and Green, N. M. (1982) FEBS Lett. 146, 87-92).

Adenosine Triphosphate↗
PubMed1984

The isolation and sequence analysis of vasoactive intestinal peptide from a ganglioneuroblastoma.

A ganglioneuroblastoma was excised at surgery from a 1-yr-old girl with severe watery diarrhea. The tumor, weighing 1 g, was extracted in trifluoracetic acid and contained 8.3 nmol immunoreactive vasoactive intestinal peptide. The peptide was isolated by affinity chromatography and high pressure liquid chromatography and was found to be identical to porcine vasoactive intestinal peptide by amino acid analysis and microsequence analysis.

Amino Acid Sequence↗
PubMed1983

Amino acid sequences of three bombesin-like peptides from canine intestine extracts.

Amino acid sequences of three canine bombesin-like peptides were determined after sequential purification of an extract obtained from 820 g of intestinal muscle. These three peptides contained 27, 23, and 10 amino acid residues. The sequences of the two shorter forms were identical to the corresponding carboxyl-terminal sequence of the heptacosapeptide. The sequence of the largest peptide is H2N-Ala-Pro-Val-Pro-Gly-Gly-Gln-Gly-Thr-Val-Leu-Asp-Lys-Met-Tyr-Pro-Arg-Gly-Asn - His(Trp-Ala-Val-Gly-His-Leu-Met-CONH2). The sequence of the 23-residue peptide is H2N-Gly-Gly-Gln-Gly-Thr-Val-Leu-Asp-Lys-Met-Tyr-Pro-Arg-Gly-Asn-His-Trp-Ala- Val-Gly(His-Leu-Met-CONH2). The sequence of the decapeptide is: H2N-Gly-Asn-His-Trp-Ala-Val-Gly-His-Leu-Met-CONH2. Comparison of the 27-residue peptide with the known structure of porcine gastrin-releasing peptide, another bombesin-like heptacosapeptide, reveals four amino acid substitutions: canine bombesin-like peptide had Pro 4, Gly 5, Gln 7, Asp 12, whereas porcine gastrin-releasing peptide had Ser 4, Val 5, Gly 7, Ala 12. Radioimmunoassay of brain extracts after similar purification revealed the presence of similar large and small forms of immunoreactive bombesin peptides, but with apparent tissue concentrations of about 4% of those present in intestinal muscle. Canine bombesin-like peptides represent another example of mammalian neuropeptides existing in more than one biologically active molecular form.

Amino Acid Sequence↗
PubMed1983

Cloning the heavy chain of human HLA-DR antigen using synthetic oligodeoxyribonucleotides as hybridization probes.

The recent development of the amino acid microsequence technique allows us to obtain partial sequence information using an extremely small amount of protein. Two sets of mixed oligonucleotide probes were chemically synthesized using the amino acid sequence information for the heavy chain of human HLA-DR antigen obtained by the microsequence technique. These two hybridization probes were used to screen cDNA clones constructed from cytoplasmic poly(A)+ mRNA from a human B lymphoblastoid homozygous cell line (LG-2). Of the 10,000 clones screened, two clones hybridized with the probes. DNA sequence analysis showed that the longer one of the two cDNA clones was 1183 nucleotides long, including the entire coding region, the signal peptide region, and the complete 3'-noncoding region. The deduced amino acid sequence of the HLA-DR alpha chain is identical to that of other cell lines with a different HLA-DR typing. However, several nucleotide differences are found in the 3'-untranslated region compared with that of other DR haplotypes.

Amino Acid Sequence↗
PubMed1983

Structure of somatostatin isolated from bovine retina.

Somatostatin-like immunoreactivity (SLI) from bovine retina was purified and its structure determined. Retinal tissue (1868 g) extracted with 3% acetic acid yielded 18.6 nmol SLI. This peptide was purified by chromatography on an affinity column made with anti-somatostatin antiserum, a reverse-phase C-18 HPLC column, and three sequential applications on a reverse-phase phenyl HPLC column. The peptide was purified 103,000-fold from the initial extract with an overall yield of 14.4%. Amino acid sequence determination by an automatic Edman degradation technique revealed the sequence to be as follows: Ser-Ala-Asn-Ser-Asn-Pro-Ala-Met- Ala-Pro-Arg-Glu-Arg-Lys-Ala-Gly-(Cys)-Lys- Asn-Phe-Phe-Trp-Lys-Thr-(Phe, Thr, Ser, Cys). The apparent identity of this peptide with somatostatin octacosapeptide (S28) purified from other mammalian tissue indicates the phylogenetic conservation of its structure and facilitates the use of the retina as a model system for studying the neurotransmitter function of somatostatin.

Amino Acid Sequence↗
PubMed1981

Unique amino terminal structure of rat little gastrin.

The heptadecapeptide form of rat gastrin was purified by a combination of DEAE cellulose, Sephadex G50 affinity, and high performance liquid chromatography. An amino terminal pyroglutamyl blocking group was removed by incubation with PCA peptidase. Amino acid analysis before and after the unblocking reaction revealed the presence of one additional residue of arginine and proline compared with porcine gastrin. Microsequencing analysis of the unblocked peptide revealed that the sequence of the remaining hexadecapeptide was RPPMEEEEEAYGWMDF. The corresponding sequence of porcine gastrin is GPWMEEEEEAYGWMDF amide. The presence of carboxyl-terminal amide group in rat gastrin is strongly supported by complete immunoreactivity with antibodies specific for amidated C-terminal sequences of mammalian gastrins. The Arg and Pro substitutions in the amino terminal region can explain poor crossreactivity of rat gastrin with antibodies specific for the amino-terminal portion of porcine or human gastrin and its more basic chromatography pattern on ion exchange resins.

Amino Acid Sequence↗
PubMed1980

Serum and tissue immune complexes in infective endocarditis.

Serial circulating immune complex (IC) determinations were performed in 24 patients with infective endocarditis (IE) using the solid phase Clq, solid phase conglutinin and 3.5% polyethylene glycol precipitation assays. Circulating IC were detected in 67% of IE patients at presentation, but in only 7% of valve lesion controls. Serial determinations produced a 75% prevalence of IC in IE. The presence of circulating IC correlated with "subacute" disease, the presence of tissue deposits of immunoglobulin and/or complement components and with certain extravalvular manifestations (immune complex type glomerulonephritis cutaneous vasculitis and musculoskeletal manifestations). Effective therapy was associated with a fall in circulating IC levels, an effect which was well demonstrated by 3 patients in whom IC rapidly fell to zero following artificial valve replacement. The results support a role for circulating IC in the pathogenesis of this disorder, and suggest that serial IC determinations are useful in following clinical progress, particularly in culture negative endocarditis.

Adult↗