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N Kalkkinen

Publications and source records attributed to N Kalkkinen.

123 records · Page 7Linked to original sources
PubMed1986

Placental protein 12 is a decidual protein that binds somatomedin and has an identical N-terminal amino acid sequence with somatomedin-binding protein from human amniotic fluid.

Placental protein 12 (PP12) was originally isolated from term human placenta and adjacent membranes. Recently we found that the site of PP12 synthesis is decidua but not placenta. In this work, the purity of PP12 was first tested by sodium dodecyl sulfate polyacrylamide slab-gel electrophoresis and by reverse phase HPLC, and the N-terminal amino acid sequence of 15 residues was determined by a liquid-phase sequencer. A single amino acid sequence of Ala-Pro-Trp-Gln-Cys-Ala-Pro-Cys-Ser-Ala-Asp-Glu-Leu-Ala-Leu was obtained showing identity to the known N-terminal amino acid sequence of somatomedin-binding protein from human amniotic fluid. Like the latter, PP12 bound somatomedin (insulin-like growth factor I) as demonstrated in gel chromatography by a shift in the elution pattern of [125I]iodo-insulin-like growth factor I after incubation with PP12. These data show that PP12 is a somatomedin-binding protein and extend through previous literature on PP12 the existing knowledge on the physiology and pathophysiology of somatomedin-binding protein(s) in human reproduction and cancer.

Amino Acid Sequence↗
PubMed1985

Identification of the major human sialoglycoprotein from red cells, glycophorin AM, as the receptor for Escherichia coli IH 11165 and characterization of the receptor site.

The pyelonephritogenic Escherichia coli strain 1 H 11165 specifically agglutinates human erythrocytes carrying the M blood group antigen. The polymorphic forms of this antigen, M and N, are located in the NH2-terminal region of the major human red-cell sialoglycoprotein, glycophorin A. Radioactively labeled glycophorin A from M cells specifically bound to the bacteria. Purified glycophorin AM, but not glycophorin AN, efficiently inhibited for binding. Mild periodate treatment oxidized the NH2-terminal serine in glycophorin AM and this resulted in loss of binding to the bacteria. High concentrations of serine and alkali-labile oligosaccharides derived from glycophorin AM inhibited the binding, whereas the synthetic M-specific NH2-terminal pentapeptide Ser-Ser-Thr-Thr-Gly did not. Neuraminidase treatment of glycophorin AM did not destroy the binding. The most efficient inhibition of binding was observed with the N-terminal glyco-octapeptide obtained from glycophorin AM by CNBr cleavage. This peptide contains both the essential serine residue and the alkali-labile oligosaccharides, which both are recognized by the bacterium.

Amino Acid Sequence↗
PubMed1984

Three genes code for rubella virus structural proteins E1, E2a, E2b and C.

The structural proteins E1, E2a, E2b and C of rubella virus (RV) were purified by preparative SDS-PAGE. The individual proteins were subjected to amino-terminal sequence analysis by Edman degradation, carboxyl-terminal structure analysis by digestion with carboxypeptidases and quantitative amino acid composition analysis. The partial amino-terminal sequences of E2a and E2b were identical and different from that of E1. The C protein did not yield any consistent results on Edman degradation, suggesting that its amino-terminus is blocked. The amino acid compositions of E2a and E2b were very similar and differed from that obtained for E1 and C, which also differed from each other. Carboxypeptidase digestions showed that E2a and E2b have an identical carboxyl-terminal structure, which differed from that of the C protein. No amino acid residues were released from the E1 protein by digestion with a mixture of carboxypeptidases A and B. These results confirm that the structural proteins of RV are translated from three genes corresponding to C, E2 and E1. E2 exists in virions in two post-translationally modified forms, E2a and E2b, which have an identical apoprotein moiety. The partial amino acid sequence information obtained here should also be sufficient to localize the ends of the individual genes on the 24S mRNA genome once its nucleotide sequence has been established.

Amino Acids↗
PubMed1983

Amino acid sequence of alpha-amylase from Bacillus amyloliquefaciens deduced from the nucleotide sequence of the cloned gene.

We have isolated by molecular cloning the gene coding for the alpha-amylase (1,4-alpha-D-glucan glucanohydrolase, EC 3.2.1.1.) from Bacillus amyloliquefaciens and determined its complete nucleotide sequence. The gene cloned in the plasmid pUB110 using Bacillus subtilis as a host, was contained in a 2.3-kilobase insert. Starting from an ATG initiator codon, an open reading frame comprising a total of 514 amino acids (1542 base pairs) was found within the cloned DNA fragment. The gene region encoding the COOH terminus of alpha-amylase was located by direct COOH-terminal analysis of the purified exoenzyme. The NH2-terminal portion of the gene encodes a 31 amino acid-long signal peptide (Palva, L., Pettersson, R. F., Kalkkinen, N., Lehtovaara, P., Sarvas, M., Söderlund, H., Takkinen, K., and Kääriäinen, L. (1981) Gene 15, 43-51). Since the signal peptide is correctly cleaved in the new host, as shown here by direct NH2-terminal sequence analysis, the exoamylase consists of 483 amino acid residues, corresponding to a molecular weight of 54,778. The reading frame used to deduce the amino acid sequence was found to be correct by comparison with partial amino acid sequence data published previously (Detera, S. D., and Friedberg, F. (1979) Int. J. Peptide Protein Res. 14, 364-372; Chung, H., and Friedberg, F. (1980) Biochem. J. 185, 387-395). Several differences between the sequence presented here and the partial ones published previously, however, were found. The nucleotide sequences both 5' and 3' to the alpha-amylase gene revealed palindromic structures including a stretch of six T-residues, suggesting transcription termination signals on both sides of the gene. Thus, it appears that alpha-amylase is translated from a monocistronic mRNA.

Amino Acid Sequence↗
PubMed1983

Rubella virus contains one capsid protein and three envelope glycoproteins, E1, E2a, and E2b.

We have analyzed the structure of rubella virus proteins labeled metabolically with [35S]methionine, [3H]mannose, and [3H]glucosamine or externally with [3H]borohydride after galactose oxidase treatment. Four structural proteins, with MrS of about 58,000 (E1), 47,000 (E2a), 42,000 (E2b), and 33,000 (C), were resolved on sodium dodecyl sulfate-polyacrylamide gels. Tryptic peptide maps obtained from [35S]methionine-labeled proteins indicated that E1 and C were unrelated to each other and to E2a and E2b, whereas the latter two gave similar, if not identical, maps. E1, E2a, and E2b were associated with the envelope and were located externally on the virus particle, whereas the C protein was associated with the RNA in the nucleocapsid. Solubilization of the virus with Triton X-100, followed by removal of the nucleocapsid and the detergent, resulted in the formation of soluble envelope protein complexes (rosettes) containing E1, E2a, and E2b. Although external labeling with [3H]borohydride and metabolic labeling with [3H]glucosamine suggested that all three proteins were glycosylated, only E1 and E2b were efficiently labeled with [3H]mannose. It is thus possible that the difference in migration between E2a and E2b is due to differences in glycosylation. Analysis by immunoprecipitation and sodium dodecyl sulfate-gel electrophoresis of intracellular [35S]methionine-labeled structural proteins synthesized in the presence and absence of tunicamycin supported the conclusion that E1 and E2 are glycoproteins. Unglycosylated E1 and E2 had an Mr of about 53,000 and 30,000, respectively.

Capsid↗
PubMed1982

Purification and characterization of a tumor-associated trypsin inhibitor from the urine of a patient with ovarian cancer.

Immunochemical studies on urine from a patient with ovarian cancer revealed the presence of a tumor-associated peptide. This peptide occurred in elevated concentrations in the urine of some patients with gynecologic cancer, in early amniotic fluid, and in some cancer tumor extracts from these patients as described previously (Stenman, U.-H., Huhtala, M.-L., Koistinen, R., and Seppälä, M. (1982) Int. J. Cancer 30, 53-47). The peptide has now been purified from the urine of a patient with ovarian cancer by gel chromatography, ion exchange chromatography, and reverse phase liquid chromatography. The amino acid composition of the peptide is: Lys (4), Arg (3), Asx (8), Thr (4), Ser (3), Glx (6), Pro (3), Gly (5), Ala (1), Val (2), Ile (3), Leu (4), Tyr (3), Phe (1), and Cys (6). These 56 amino acids correspond to Mr = 62000 for the peptide, a value that is in agreement with the molecular weight established by gel chromatography. The molecule contains no carbohydrate. It is microheterogeneous in charge, the isoelectric point of the main component being 5.8. The purity of the peptide was confirmed by determination of the NH2-terminal amino acid sequence. The 21 residues determined were found to be identical with the corresponding ones of human pancreatic trypsin inhibitor. The purified peptide also inhibited bovine trypsin effectively.

Amino Acid Sequence↗
PubMed1981

Nucleotide sequence of the promoter and NH2-terminal signal peptide region of the alpha-amylase gene from Bacillus amyloliquefaciens.

We have isolated and partially sequenced the gene coding for alpha-amylase (EC 3.2.1.1) from Bacillus amyloliquefaciens by molecular cloning in the plasmid pUB110 using Bacillus subtilis as a host. The nucleotide sequence of the NH2-terminal region of the cloned gene was determined and found to contain a 31-residue-long stretch of amino acids preceding the NH2-terminal sequence of the extracellular alpha-amylase. Within this sequence there is a 15-residue-long stretch of uncharged amino acids similar to that found at the NH2 terminus of other precursors to exported proteins. This "signal sequence" is probably removed in conjunction with the translocation of alpha-amylase through the cytoplasmic membrane. In vitro labeling of alpha-amylase with radioactive amino acids in a coupled transcription-translation system followed by partial sequencing established the exact location of the NH2 terminus of the alpha-amylase gene. The nucleotide sequence preceding the NH2 terminus has properties resembling the RNA-polymerase- and ribosome-binding sites found at the 5' terminus of many prokaryotic genes.

Amylases↗
PubMed1980

Analysis of Semliki-Forest-virus structural proteins to illustrate polyprotein processing of alpha viruses.

The four structural proteins of Semliki Forest virus were purified in an amount of 30-50 nmol by preparative sodium dodecylsulfate/polyacrylamide gel electrophoresis. Each protein was subjected to N-terminal structural analysis by degradation in a liquid-phase sequencer. About 20 residues were determined for each of the two membrane glycoproteins E1 and E2. The amino acid sequence of E1 but not that of E2 showed extensive homology to the corresponding proteins of the closely related Sindbis virus. Both E1 and E2 seem to lack a signal sequence at the N terminus, since the proportion of polar amino acids in this region deviates from the proportion in the known hydrophobic signal sequences. The envelope glycoprotein E3 and the capsid protein did not yield any significant result on Edman degradation, suggesting that they have blocked N-terminal amino groups.

Amino Acid Sequence↗
PubMed1977

Pseudomonas cytochrome c peroxidase. XIII. pH-denaturation of the enzyme.

The effect of pH on the oxidized Pseudomonas cytochrome c peroxidase molecule was studied by measuring the peroxidatic activity, the sedimentation velocity, the circular dichroic spectra in the far UV and Soret regions, and the optical absorption spectra of the enzyme in the pH range 2.5-13.0 at a constant ionic strength (micron = 0.1). The enzyme was stable in a narrow pH region, pH 6.0 - 7.4. In the low pH range the gross tertiary structure was observed to change quite simultaneously with the enzymatic activity and secondary structure. The optical absorption spectra indicated that there were no coordinated internal protein liqands in the 6th coordination positions of the heme prosthetic groups at the lowest pH studied. In the high pH range the secondary structure and the protein environment of hemes were observed to remain stable after the tertiary structure had changed and the activity had decreased. According to the optical absorption spectra the 6th internal protein ligands of hemes were retained at the highest pH studied.

Circular Dichroism↗
PubMedDate not supplied

Molecular and antigenic characteristics and synthesis of rubella virus structural proteins.

The molecular and antigenic properties and synthesis of the structural proteins as well as the virus-specific RNAs of rubella virus were analyzed. Virions contain three major polypeptides--E1 (relative molecular weight [Mr] 58,000), E2 (Mr 42,000-47,000), and C (Mr 33,000). E1 and E2 are glycosylated and located externally on the viral membrane. C is associated with the genomic RNA to form the nucleocapsid. E2 occurs in two forms, E2a and E2b; the protein moieties of the two are indistinguishable. E1 is the viral hemagglutinin. IgG antibodies react with all the structural proteins, whereas IgM and IgA antibodies react predominantly with E1 and C proteins, respectively. After rubella vaccination, the reactivity of IgG antibodies matured slowly and reactivity of IgM and IgA antibodies remained at a low level. Rubella virus contains a 40S (Mr approximately 3.8 X 10(6)) single-stranded RNA with a 5' cap structure and a 3' poly(A) tract. In infected cells a 24S (Mr approximately 1.2 X 10(6)) subgenomic, polyadenylated mRNA is synthesized; it codes for a precursor (Mr 110,000 [p110]) to the structural proteins. The gene order in the 24S RNA is NH2-C-E2-E1-COOH. The overall structure and strategy of gene expression of rubella virus appears to be similar to that of the alphaviruses of the Togaviridae family.

Adolescent↗