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H Kratzin

Publications and source records attributed to H Kratzin.

At least 37 records · Page 2Linked to original sources
PubMed1988

Cloning of a full-length complementary DNA for fatty-acid-binding protein from bovine heart.

A full-length cDNA for bovine heart fatty-acid-binding protein (H-FABP) was cloned from a lambda gt11 cDNA library established from bovine heart muscle. The cDNA sequence shows an open reading frame coding for a protein with 133 amino acids. Colinearity with the amino acid sequences of four tryptic peptides was asserted. H-FABP isolated from bovine heart begins with an N-acetylated valine residue, however, as derived from analysis of the tryptic, amino-terminal-blocked peptide and the molecular mass of the peptide obtained via secondary-ion mass spectrometry. The molecular mass of the total protein is 14673 Da. Bovine H-FABP is 89% homologous to rat H-FABP and 97% homologous to the bovine mammary-derived growth-inhibition factor described recently by Böhmer et al. [J. Biol. Chem. 262, 15137-15143 (1987)]. Significant homologies were also found with bovine myelin protein P2 and murine adipocyte protein p422. Secondary-structure predictions were proposed for these proteins, based on computer analysis, which reveal striking similarities.

Acetylation↗
PubMed1988

Murine Fc gamma receptor proteins: identification of a previously unrecognized molecule with a monoclonal antibody (12-15).

Previously we studied differential expression of cell surface molecules between the metastatic murine lymphoma ESb and an adhesion variant ESb-MP. Here we describe the specificity of a monoclonal antibody (12-15) that showed strong binding to the adhesion variant and weak reactivity against ESb cells. The antibody also reacted to lymphoid but not to macrophage-derived cell lines and immunoprecipitated a molecule of approx. 60-69 kDa from ESb-MP cells. N-terminal sequencing of the antigen revealed identity to the beta protein of mouse Fc gamma receptors. Using monoclonal antibodies against Fc gamma receptors (2.4G2 and K9.361) in immunofluorescence assays and cDNA probes specific for alpha, beta 1 and beta 2 Fc receptor transcripts in Northern blot experiments the differential expression of Fc receptors in ESb and ESb-MP cells was confirmed. Biochemical analysis of endoglycosidase F-treated precipitates revealed that antibody 12-15 reacted to products of all three transcripts with molecular masses for the protein core of 38.5 kDa (beta 1), 34 kDa (beta 2) and 31 kDa alpha). In addition, an unknown protein of 37 kDa (termed beta 3) was identified by antibody 12-15 which could also be detected in ESb cells and EL4 cells. Antibodies 2.4G2 and K9.361 did not react to the beta 3 chain but reacted to varying extents to the other Fc proteins in macrophage and lymphoid cells. Comparison by peptide mapping of the novel beta 3 chain to beta 1, beta 2 and alpha proteins revealed similar, but also distinct peptides. The tissue-specific reactivity of monoclonal antibody 12-15 is likely to be due to a carbohydrate epitope associated with all Fc gamma receptors in lymphoid but not macrophage cell lines.

Animals↗
PubMed1988

Isolation and primary structure of tumor-derived peptides related to human pancreastatin and chromogranin A.

Using an antiserum raised against a synthetic C-terminal peptide of porcine pancreastatin, we detected pancreastatin-like immunoreactivity (PLI) in human pancreatic islets, adrenal medulla, and endocrine tumors. From a carcinoid liver metastasis, human PLI was extracted and purified by HPLC. Two C-terminally amidated peptides were isolated and characterized by sequence analysis. The first peptide, hCgA-210-301, consists of 92 amino acid residues with glycinamide as C terminus. It is identical to the cDNA-derived sequence of human chromogranin A, positions 210-301, which is preceded by two basic residues indicating a putative processing site. The C-terminal part, positions 250-301, shows 70% sequence identity to porcine pancreastatin and represents the human pancreastatin-like sequence. The second peptide, hCgA-273-301, represents a C-terminally amidated fragment of the human pancreastatin sequence, generated by an Asp-Pro cleavage at the N terminus. Peptide hCgA-273-301 was synthesized to confirm the structure of the natural peptide. Two other peptides derived from human chromogranin A were isolated and partially characterized. They are generated by proteolytic cleavage after dibasic amino acids Lys-Arg (positions 338-339) and after Trp-376 of the human chromogranin A sequence, respectively. These results indicate that chromogranin A may represent the precursor for pancreastatin-related and possibly other yet-unidentified peptides of unknown physiological function.

Amino Acid Sequence↗
PubMed1987

Structural relationship of an apolipoprotein (a) phenotype (570 kDa) to plasminogen: homologous kringle domains are linked by carbohydrate-rich regions.

At least six allelic forms of apolipoprotein(a), differing in molecular mass, could be detected by immunoblot analysis. One of these phenotypes with a molecular mass of 570 kDa has been investigated. After reduction and carboxymethylation it was digested with trypsin and the resulting peptides were separated by gel filtration and reverse phase HPLC. The tryptic fragments sequenced comprised a total of 356 amino acids. The N-terminus of apo(a) was highly homologous to the start of the kringle 4 domain from human plasminogen and the majority of the tryptic peptides isolated was also homologous to sequences from this kringle. At least five homologous "kringle 4" domains are present in apolipoprotein(a) whereby one domain occurs more frequently than the others. A carbohydrate-rich peptide was also obtained in high yield. This glycopeptide connects two "kringle 4" domains and contains one N-glycoside within the kringle and six potential O-glycosides in the linking region. From the recovery it can be estimated that this peptide occurs several times within the whole apolipoprotein (a) sequence. The high carbohydrate content is in sharp contrast to that of human plasminogen. Other peptides sequenced indicate that apo (a) also contains domains homologous to the kringle 5 and protease regions of plasminogen. No unique peptides were found. These studies suggest that apolipoprotein (a) could have arisen through duplication of specific regions from the human plasminogen gene. The size heterogeneity of apo (a) might then be explained by differences in the numbers of gene duplications.

Amino Acid Sequence↗
PubMed1987

Structure of the yeast isoleucyl-tRNA synthetase gene (ILS1). DNA-sequence, amino-acid sequence of proteolytic peptides of the enzyme and comparison of the structure to those of other known aminoacyl-tRNA synthetases.

The ILS1 gene encoding for cytoplasmic isoleucyl-tRNA synthetase from Saccharomyces cerevisiae was subcloned from a 5.4-kb insert of the shuttle vector YEp13 to M13mp8 and M13mp9. Nucleotide sequence analysis of a 4.3-kb BamHI-HpaI fragment revealed a single open reading frame from which we deduced the amino-acid sequence of the enzyme. Independently obtained amino-acid sequence information from ten tryptic peptides of the purified enzyme confirmed the gene-derived structure. The enzyme is comprised of 1073 amino-acids consistent with earlier determinations of its molecular mass. The codon usage of ILS1 is typical of abundant yeast proteins. A significant homology to E. coli isoleucyl- and valyl-tRNA synthetases as well as to yeast valyl-tRNA synthetase was detected. The characteristic amino-acid residues of the aminoacyl-adenylate site and of the potential binding site of the 3'-end of tRNA found in other synthetases are present in the structure.

Amino Acid Sequence↗
PubMed1985

Isolation and characterization of proSS1-32, a peptide derived from the N-terminal region of porcine preprosomatostatin.

A peptide derived from the N-terminal region of porcine prosomatostatin, proSS1-32, has been purified to homogeneity from extracts of porcine upper intestine. Amino acid analysis revealed that the peptide consists of 32 residues. The complete primary structure was determined as: A P S D P R L R Q F L Q K S L A A A A G K Q E L A K Y F L A E L. This sequence obviously comprises residues 1-32 of porcine prosomatostatin since it is identical to the corresponding sequence in human preprosomatostatin. The postulated cleavage site in porcine prosomatostatin is a Leu-Leu bond between residues 32 and 33, thus confirming previous studies of the processing of the somatostatin precursor in the rat and transgenic mouse.

Amino Acid Sequence↗
PubMed1985

Valosin: isolation and characterization of a novel peptide from porcine intestine.

The isolation and primary structure of a novel gastrointestinal peptide, designated valosin, is described. The peptide was purified from porcine upper gut extracts using an HPLC and N-terminal sequence screening strategy which depends on chromatographic and structural characteristics as isolation criterion. The amino acid sequence of this peptide consists of 25 amino acid residues:

Amino Acid Sequence↗
PubMed1984

[Primary structure of human class II histocompatibility antigens (HLA-D). I. Isolation, purification and characterization of the HLA-D alpha/beta chain complex from a homozygous lymphoblastoid B cell line, H2LCL (HLA-A3,3;B7,7;Dw2, 2;DR2,2;MT1,1;DC1,1;MB1,1].

The complex of alpha and beta chains of HLA-D membrane antigens has been isolated from a lymphoblastoid homozygous B cell line, H2LCL (HLA-A3,3; B7,7; Dw2,2; DR2,2; MT1,1; DC1,1; MB1,1), by an exclusively chemical two-step procedure and characterized by electrophoresis as well as isoelectric focusing in polyacrylamide gel. Cells were gained using long term cultivation in large scale, the crude membrane by differential centrifugation. The proteins of the crude membrane were then solubilized in NP-40, pH 5.0. The first purification step for HLA-D antigens consisted in an ion-exchange chromatography on carboxymethyl cellulose using the solubilization buffer. By this procedure the complex of proteins with relative molecular masses of Mr approximately 34 000 and Mr approximately 29 000 was in a high percentage not bound to the carboxymethyl cellulose. The bound fraction contained the HLA-A, -B and -C antigens and a component with Mr approximately 31 000 corresponding to the well known Ii-fraction. The bound proteins could be recovered from the column by a sodium chloride gradient. The proteins not bound to the carboxymethyl cellulose were precipitated with acetone, dissolved, dialysed against SDS buffer, pH 7.2 and then submitted to the second purification step, the Sephacryl S-300 chromatography. By this procedure the corresponding complex could be further separated from higher and lower molecular proteins. The complex was used as the starting material for the separation of alpha and beta chains. Amino-acid sequences established of the isolated chains have already been communicated.

Amino Acid Sequence↗
PubMed1984

[Primary structure of human class II histocompatibility antigens (HLA-D). II. Separation of alpha from beta-chains of HLA-D from a homozygous lymphoblastoid B cell line, H2LCL (HLA-A3,3;B7,7;Dw2,2;MT1,1;DC1,1;MB1,1), and characterization of the isolated chain].

In a previous communication we described the preparation of the alpha/beta-chain complex of HLA-D membrane antigens from a homozygous lymphoblastoid B cell line, H2LCL (HLA-A3,3; B7,7; Dw2,2; DR2,2; MT1,1; DC1,1; MB1,1). In this paper we present the separation of the alpha- and beta-chains in quantitative scale using hydroxylapatite chromatography with a pH 7.15 phosphate buffer, containing sodium dodecyl sulfate. An alternative isolation procedure is electrophoresis, taking advantage of the different isoelectric points of the alpha- and beta-chains. The relative molecular masses, the isoelectric points and the N-terminal sequences are discussed and compared with the results of other investigators. Remarkable is the polymorphism of both the chains, especially the beta-chains. The importance of the previously described three step purification procedure for the preparation of these antigens for sequence studies has been pointed out.

B-Lymphocytes↗
PubMed1983

Primary structure of human class II histocompatibility antigens 3rd communication. Amino acid sequence comparison between DR and DC subclass antigens derived from a lymphoblastoid B cell line homozygous at the HLA loci (HLA-A3,3; B7,7; Dw2,2; DR2,2: MT1,1; Dc1,1: MB1,1).

Class II histocompatibility antigens from a homozygous human lymphoblastoid B cell line (HLA-A3,3; B7,7; Dw2,2; DR2,2; MT1,1; DC1,1; MB1,1) were isolated by an exclusively chemical procedure. After the separation of the alpha and beta chains the amino acid sequences of the extracellular parts of the DR2 alpha and the DR2 beta chains could be determined, in spite of the heterogeneity of both fractions. Peptides isolated in considerably lower yields indicated at least one more alpha chain and 6 further beta chains. By the investigation of these lower-yield enzymatic fragments, in addition to the DR2 alpha and DR2 beta chains, we can now report on the partial amino acid sequences of a DC alpha chain, a second beta chain of DR type and one beta chain of DC type. Furthermore, the isolation of a characteristic glycopeptide suggests the existence of a second beta chain of DC type. The following class II antigens are therefore identified: DR2 alpha/DR2 beta 1, DR2 alpha/DR2 beta 2, DC1 alpha/DC1 beta 1, DC1 alpha/DC1 beta 2, indicating that each subclass can be subdivided into two antigens because each alpha chain type can associate with at least either beta 1 or beta 2. Further investigations must clarify whether the remaining three beta chains are of the DR, DC, or of a third beta chain type. Amino acid sequence comparisons confirm that the human DR antigens are the equivalent of mouse I-E antigens, while human DC antigens are the equivalent of mouse I-A antigens.

Amino Acid Sequence↗
PubMed1982

[Primary structure of class II human histocompatibility antigens. 2nd Communication. Amino acid sequence of the N-terminal 179 residues of the alpha-chain of an HLA-Dw2/DR2 alloantigen (author's transl)].

From a lymphoblastoid homozygous cell-line (HLA-A3,3; B7,7; Dw2,2; DR2,2) the alpha-chain of the HLA-Dw2/DR2 antigen was isolated by an exclusively chemical procedure. The alpha- was separated from the beta-chain by chromatography with hydroxylapatite in Na-dodecyl sulfate. Here we describe the amino acid sequence of the alpha-chain up to Position 179. The molecule is divided into two domains which do not appear homologous to each other but show a significant homology to the beta-chain (21.2%). The similarity is larger in the second (27%) than in the first (15.5%) domain, indicating a different evolutionary relationship for both parts. In contrast to the beta-chain both domains contain an N-glycosidically linked carbohydrate. The methionines are positioned only in the N-terminal, the cysteines exclusively in the C-terminal domain. Only the latter can therefore be stabilized by a disulfide bridge. As with the beta-chain the regions around the cysteines show a remarkable similarity with the constant C-terminal domains of k-, lambda-, alpha-, gamma-, sigma-, epsilon- and mu-chains of immunoglobulins. Although to a considerably lesser extent, the alpha-chain preparation also shows a heterogeneity at the protein level. Since the employed cell-line is homozygous with regard to HLA-D/DR, our results indicate that at least two alpha-chain genes exist in the HLA-D/DR-region. Together with the already published sequence of the beta-chain, the extracellular part of an histocompatibility antigen of the HLA-D type is now known.

Amino Acid Sequence↗
PubMed1981

[Chromatography and rechromatography in high-performance liquid chromatography of peptide mixtures: the complete primary structure of an immunoglobulin L-chain of kappa-type, subgroup I (Bence-Jones Protein Den) (author's transl)].

In continuation of our work on the separation of enzymatic hydrolysates by high-pressure liquid chromatography with reverse-phases we present two new buffer systems: firstly 0.005M potassium phosphate, pH 6.0 and secondly aqueous trifluoroacetic acid, pH 2.15. As organic solvent acetonitrile is always used. The excellent properties are demonstrated by the separation of the tryptic peptides of Bence-Jones protein Den (Mr = 23000). Like ammoniumacetate these buffers are well suited for the first separation of peptide mixtures but can be used rather effectively in rechromatographies. If peaks are not or not fully resolved during the first chromatography they can be separated by a rechromatography in one of the other systems. This was most successful since in both cases the same high resolving technique is employed. From the primary separation 14 peptides could be isolated in analytically pure form. The remaining fragments were completely purified after one rechromatography. The amino acid analysis yielded also integer numbers and by a modified Edman degradation the primary structure could be determined. As with protein Wes (Kratzin, H., Yang, C.Y., Krusche, T.U. & Hilschmann, N. (1980) Hoppe-Seyler's Z. Physiol. Chem. 361, 1591--1598) all the peptides were recovered after high-pressure liquid chromatography, none was missing. From these data and from homology reasons the complete amino acid sequence of protein Den could be established. It contains 214 residues and belongs to subgroup I of the kappa-chains. The valine residue in position 191 indicates that it belongs to allotype Inv b+.

Amino Acid Sequence↗
PubMed1981

[Primary structure of class II human histocompatibility antigens. 1st communication. Amino acid sequence of the N-terminal 198 residues of the beta chain of a HLA-Dw2,2;DR2,2-alloantigen (author's transl)].

The beta-chains of HLA-D/DR antigens from a lymphoblastoid homozygous cell line (HLA-A3,3;B7,7;-Dw2,2;DR2,2) were purified by an exclusively chemical three step procedure: 1) Nonidet P-40/CM-cellulose ion-exchange-, 2) Na-dodecyl sulfate/sephacryl S-300- and 3) Na-dodecyl sulfate/hydroxylapatite chromatography. The amino acid sequence of the beta chain was established up to position 198. These residues comprise the extra-cellular part of the molecule which consists of two domains each probably containing a disulfide bridge, like immunoglobulin L chains. Within the first domain the carbohydrate moiety is attached to asparagine in position 19. Sequence work was hampered by the fact that the beta chain preparation of the HLA-Dw2,2;DR2,2 antigen actually consists of a pool of beta chains. The beta chain, the sequence of which has now been established, represents about 70% of the beta chain pool. The remaining 30% consist of at least 6 more chains which are homologous among one another as well as to the described beta chain. Since the antigens were gained from a homozygous lymphoblastoid cell-line our results indicate that the HLA-D region codes for at least 7 beta-chains and must therefore consist of a cluster of beta chain genes.

Amino Acid Sequence↗
PubMed1980

[Preparative separation of the tryptic hydrolysate of a protein by high-pressure liquid chromatography. The primary structure of a monoclonal L-chain of k-type, subgroup I (Bence-Jones Protein Wes) (author's transl)].

The tryptic hydrolysate of Bence-Jones protein Wes (Mr = 23000) was separated by high-pressure liquid chromatography in a volatile buffer system on a reversed-phase column. From the resulting 22 peptides 18 yielded integer numbers after amino acid analysis. They could be used directly for a modified Edman degradation. Two peptides were not separated by this procedure, two were missing. Under different conditions these four peptides could also be prepared in pure form. 3 mg was sufficient to elucidate the primary structure of the variable part of this protein completely. The arrangement of the tryptic peptides was deduced by homology to other k=chains. Protein Wes contains 214 residues and belongs to subgroup I of the k-chains. The valine residue in position 191 indicates that it belongs to allotype Inv b+.

Bence Jones Protein↗
PubMed1979

[Rule of antibody structure. Primary structure of a human monoclonal IgA-immunoglobulin (myeloma protein Tro). VII. Purification and characterization of the disulfide bridges].

Myeloma Protein Tro has been isolated from the plasma of a myeloma patient. Monomeric IgA was separated from its polymer (by chromatography on Sephadex G-200). Both the forms were split with pepsin or cyanogen bromide and, if necessary, with thermolysin and subtilisin. The cystin-containing peptides were isolated from the hydrolysates by chromatography on Sephadex, ion-exchange columns, preparative paper chromatography, thin-layer chromatography, electrophoresis or by a combination of these methods. They were characterized by amino acid analyses and by determination of the N-terminal amino acids using the Dansyl-Edman procedure. Thus all the disulfide bridges of an IgA1 immunoglobulin could be established. The monomer has all together 48 cysteins, seven in each L- and seventeen in each H-chain; all these are covalently bonded by SS-bridges. Free SH-groups were not detected. The J-chain could only be identified serologically in the polymeric form of the protein. It is shown that the subunits of the polymers are covalently attached through either Cysl3, Cysl7 or both these residues of the H-chain.

Amino Acid Sequence↗
PubMed1978

Genetic determination of antibody specificity. Gene translocation and fusion, the molecular basis for the differentiation of the antibody-producing cell.

The best system for the study of cell differentiation is a cell which in its differentiated state differs only by one product. This is the case in the immune system. The undifferentiated, but omnipotent stem cell differentiates into a committed B cell which produces only one type of specific antibody out of a million different, genetically fixed possibilities. Gene translocation and fusion is the basis of this differentiation process.

Alleles↗