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  • 1. Ahmed, A. Ahmed
    et al.
    El-Seedi, Hesham R.
    Mahmoud, Ahmed A.
    El-Douski, Abd El-Aziz A.
    Zeid, Ibrahim F.
    Bohlin, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Eudesmane derivatives from Laggera crispata and Pluchea carolonesis1998Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 49, nr 8, s. 2421-2424Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Investigation of the aerial parts of Laggera crispata and Pluchea carolonesis afforded in addition to several known compounds, three new eudesmane derivatives, 3β,4α-dihydroxy-7-epi-eudesm-11(13)-ene, 3α-(2′,3′-dihydroxy-2′-methylbutanoyl)-4,11-dihydroxy-6,7-dehydroeudesman-8-one and 3α-(3′-chloro-2′-hydroxy-2′-methylbutanoyl)cuauhtemone. The structures were elucidated by spectroscopic methods

  • 2. Andersson, Derek
    et al.
    Chakrabarty, Romit
    Bejai, Sarosh
    Zhang, Jiaming
    Rask, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinsk biokemi och mikrobiologi.
    Meijer, Johan
    Myrosinases from root and leaves of Arabidopsis thaliana have different catalytic properties2009Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 70, nr 11-12, s. 1345-1354Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Myrosinases (EC 3.2.1.147) are beta-thioglucoside glucosidases present in Brassicaceae plants. These enzymes serve to protect plants against pathogens and insect pests by initiating breakdown of the secondary metabolites glucosinolates into toxic products. Several forms of myrosinases are present in plants but the properties and role of different isoenzymes are not well understood. The dicot plant model organism Arabidopsis thaliana seems to contain six myrosinase genes (TGG1-TGG6). In order to compare the different myrosinases, cDNAs corresponding to TGG1 from leaves and TGG4 and TGG5 from roots were cloned and overexpressed in Pichia pastoris. The His-tagged recombinant proteins were purified using affinity chromatography and the preparations were homogenous according to SDS-PAGE analysis. Myrosinase activity was confirmed for all forms and compared with respect to catalytic activity towards the allyl-glucosinolate sinigrin. There was a 22-fold difference in basal activity among the myrosinases. The enzymes were active in a broad pH range, are rather thermostable and active in a wide range of salt concentrations but sensitive to high salt concentrations. The myrosinases showed different activation-inhibition responses towards ascorbic acid with maximal activity around 0.7-1 mM. No activity was registered towards desulphosinigrin and this compound did not inhibit myrosinase activity towards sinigrin. All myrosinases also displayed O-beta-glucosidase activity, although with lower efficiency compared to the myrosinase activity. The differences in catalytic properties among myrosinase isozymes for function in planta are discussed.

  • 3.
    Andersson Dunstan, Christina
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Liu, Boling
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Welch, Christopher J
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för organisk farmaceutisk kemi.
    Perera, Premila
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Bohlin, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Alphitol, a phenolic substance from Alphitonia zizyphoides which inhibits prostaglandin biosynthesis in vitro1998Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 48, nr 3, s. 495-497Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The new phenolic compound, 3,5-dihydroxy-4-methoxy phenethyl alcohol, named alphitol, and betulinic acid were isolated from the bark of Alphitonia zizyphoides. The chemical structure of alphitol was determined by mass spectrometry in combination with one and two dimensional NMR, including HMBC. Both compounds inhibited prostaglandin biosynthesis in vitro, alphitol with an IC50 value of 0.66 mM, which is of the same magnitude as acetyl salicylic acid.

  • 4. Broussalis, Adriana M.
    et al.
    Göransson, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Coussio, Jorge D.
    Ferraro, Graciela
    Martino, Virginia
    Claeson, Per
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    First cyclotide from Hybanthus (Violaceae)2001Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 58, nr 1, s. 47-51Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Hypa A, a novel macrocyclic polypeptide containing 30 amino acid residues, has been isolated from the n-butanol extract of the Argentine plant Hybanthus parviflorus. The sequence, cyclo-(SCVYIPCTITALLGCSCKNKVCYNGIPCAE), was determined by automated Edman degradation, quantitative amino acid analysis and nanospray MS/MS(2). Three intramolecular disulfide bridges stabilize the cyclic peptide backbone of hypa A. Using these structural features to classify the peptide as a cyclotide, we extended the distribution of that substance class to a new genus, and now propose a uniform nomenclature for cyclotides.

  • 5. Broussalis, Adriana M.
    et al.
    Göransson, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Coussio, Jorge D.
    Ferraro, Graciela
    Martino, Virginia
    Claeson, Per
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    First cyclotide from Hybanthus (Violaceae)2001Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 58, nr 1, s. 47-51Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Hypa A, a novel macrocyclic polypeptide containing 30 amino acid residues, has been isolated from the n-butanol extract of the Argentine plant Hybanthus parviflorus. The sequence, cyclo-(SCVYIPCTITALLGCSCKNKVCYNGIPCAE), was determined by automated Edman degradation, quantitative amino acid analysis and nanospray MS/MS2. Three intramolecular disulfide bridges stabilize the cyclic peptide backbone of hypa A. Using these structural features to classify the peptide as a cyclotide, we extended the distribution of that substance class to a new genus, and now propose a uniform nomenclature for cyclotides.

  • 6.
    Burman, Robert
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Gruber, Christian W
    Rizzardi, Kristina
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Herrmann, Anders
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Craik, David J
    Gupta, Mahabir P
    Göransson, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Cyclotide proteins and precursors from the genus Gloeospermum: filling a blank spot in the cyclotide map of Violaceae2010Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 71, nr 1, s. 13-20Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Cyclotides are disulfide-rich plant proteins that are exceptional in their cyclic structure; their N and C termini are joined by a peptide bond, forming a continuous circular backbone, which is reinforced by three interlocked disulfide bonds. Cyclotides have been found mainly in the coffee (Rubiaceae) and violet (Violaceae) plant families. Within the Violaceae, cyclotides seem to be widely distributed, but the cyclotide complements of the vast majority of Violaceae species have not yet been explored. This study provides insight into cyclotide occurrence, diversity and biosynthesis in the Violaceae, by identifying mature cyclotide proteins, their precursors and enzymes putatively involved in their biosynthesis in the tribe Rinoreeae and the genus Gloeospermum. Twelve cyclotides from two Panamanian species, Gloeospermum pauciflorum Hekking and Gloeospermum blakeanum (Standl.) Hekking (designated Glopa A-E and Globa A-G, respectively) were characterised through cDNA screening and protein isolation. Screening of cDNA for the oxidative folding enzymes protein-disulfide isomerase (PDI) and thioredoxin (TRX) resulted in positive hits in both species. These enzymes have demonstrated roles in oxidative folding of cyclotides in Rubiaceae, and results presented here indicate that Violaceae plants have evolved similar mechanisms of cyclotide biosynthesis. We also describe PDI and TRX sequences from a third cyclotide-expressing Violaceae species, Viola biflora L., which further support this hypothesis.

  • 7.
    El-Seedi, Hesham R
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Gohil, Suresh
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Kemiska sektionen, Kemiska institutionen.
    Perera, Premila
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Torssell, Kurt B G
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Bohlin, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Cyclopeptide alkaloids from Heisteria nitida1999Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 52, nr 8, s. 1739-1744Artikel i tidskrift (Refereegranskat)
  • 8.
    Henz Ryen, Astrid
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Farmakognosi.
    Backlund, Anders
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Farmakognosi.
    Kogej, Thierry
    Structural classification and scaffold diversity of sesquiterpene lactones in the angiospermsIngår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700Artikel i tidskrift (Övrigt vetenskapligt)
    Abstract [en]

    Sesquiterpene lactones (STLs) present one of the largest groups of plant specialized metabolites with a wide range of biological activities. They are a valuable source for new plant derived drugs and drug leads since they contain several important chemical properties responsible for their versatile therapeutic potential.

    The aim of this study was to analyze and compare the chemical diversity of all types of STLs in different plant groups, both qualitatively and quantitatively. For this purpose, over 5,200 STLs have been compiled and their plant origin has been recorded, resulting in a comprehensive dataset comprising over 8,600 entries. An overview of skeleton classes and their distribution among plant families was given by assigning the STLs to their major classes. An extensive scaffold diversity analysis was performed based on the molecular framework of these compounds using established metrics. Furthermore, molecular diversity and similarity was assessed via 2D fingerprint and clustering analysis.

    The results highlighted significant differences in the degree of chemical diversity. It was demonstrated that the investigated plant families have tendencies to produce certain types of skeletons. The quantity and distribution of skeleton classes was determined per plant family and genus, as well as the proportions of skeleton classes to other STL producing families. Analyzing the scaffold diversity showed that they possessed specific sets of molecular frameworks with a considerable variation in their frequency of occurrence. Even if many plant families produce STLs belonging to the same skeleton class, their corresponding molecular frameworks differ. Clustering analysis confirmed the known large structural diversity and revealed similarities and differences of the compounds. The metrics employed enabled to qualitatively divide STLs into smaller groups with similar structural features, which reflected biologically and chemically different STLs and pointed out the differentiation of various plant groups, down to the taxonomic rank of the species.

    Taken together, these analyses provided a comprehensive insight into scaffold and molecular diversity of STLs. Due to the detailed taxonomic annotation, the distinct distribution of different types of STLs was captured. This dataset represents the latest detailed compilation of STLs in the angiosperms, which can be used as a basis for further chemoinformatic or chemosystematic analyses. To provide an example of potential implementations, the results were utilized in a phylogenetic exploration of these metabolites.

  • 9.
    Herrmann, Anders
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Burman, Robert
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Mylne, Joshua S.
    Karlsson, Gustav
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Gullbo, Joachim
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska fakulteten, Institutionen för medicinska vetenskaper, Klinisk farmakologi.
    Craik, David
    Clark, Richard
    Göransson, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    The alpine violet, Viola biflora, is a rich source of novel cyclotides with potent cytotoxic cytotoxicity2008Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 69, nr 4, s. 939-952Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    The cyclotides are currently the largest known family of head-to-tail cyclic proteins. The complex structure of these small plant proteins, which consist of approximately 30 amino acid residues, contains both a circular peptide backbone and a cystine knot, the combination of which produces the cyclic cystine knot motif. To date, cyclotides have been found in plants from the Rubiaceae, Violaceace and Cucurbitaceae families, and are believed to be part of the host defence system. In addition to their insecticidal effect, cyclotides have also been shown to be cytotoxic, anti-HIV, antimicrobial and haemolytic agents. In this study, we show that the alpine violet Viola biflora (Violaceae) is a rich source of cyclotides. The sequences of 11 cyclotides, vibi A-K, were determined by isolation and MS/MS sequencing of proteins and screening of a cDNA library of V. biflora in parallel. For the cDNA screening, a degenerate primer against a conserved (AAFALPA) motif in the cyclotide precursor ER signal sequence yielded a series of predicted cyclotide sequences that were correlated to those of the isolated proteins. There was an apparent discrepancy between the results of the two strategies as only one of the isolated proteins could be identified as a cDNA clone. Finally, to correlate amino acid sequence to cytotoxic potency, vibi D, E, G and H were analysed using a fluorometric microculture cytotoxicity assay using a lymphoma cell line. The IC50-values of the bracelet cyclotides vibi E, G and H ranged between 0.96 and 5.0 mu M while the Mobius cyclotide vibi D was not cytotoxic at 30 mu M.

  • 10.
    Ibewuike, Jospeh C
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Ogundaini, Abiodun O
    Ogungbamila, Francis O
    Martin, Marie-Thérese
    Gallard, Jean-Francois
    Bohlin, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Païs, Mary
    Piliostigmin, a 2-phenoxychromone, and C-methylflavonols from Piliostigma thonningii1996Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 43, nr 3, s. 687-690Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Piliostigmin, a 2-phenoxychromone, and three C-methylflavonols, 6,8-di-C-methylquercetin 3-methyl ether, 6-C-methylquercetin 3,7-dimethyl ether and 6,8-di-C-methylquercetin 3,7-dimethyl ether, were isolated from the leaves of Piliostigma thonningii, together with the known compounds quercetin, quercitrin, 6-C-methylquercetin S-methyl ether, 6-C-methylquercetin 3,7,3'-trimethyl ether, 6,8-di-C-methyllraempferol 3-methyl ether and 6,8-di-C-methyllraempferol 3,7-dimethyl ether. The structures of the new compounds were established by spectral methods, especially 2D NMR. Complete C-13 assignments using HMBC experiments are reported for the four latter C-methylflavonols.

  • 11.
    Larsson, Sonny
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi.
    The "new" chemosystematics: Phylogeny and phytochemistry2007Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 68, nr 22-24, s. 2904-2908Artikel, forskningsöversikt (Refereegranskat)
    Abstract [en]

    For almost a decade it has been acknowledged that the flowering plant dichotomy of monocotyledons and dicotyledons does not reflect the evolution of angiosperms. Despite this, conclusions in the field of chemosystematics are still drawn from, and rely on, non-phylogenetic botanical classifications such as those of Cronquist, Dahlgren and Takhtajan. In this paper the two alkaloids colchicine and camptothecin are used as examples of how phylogenetic systematics may be applied to alkaloid chemosystematics.

  • 12.
    Slazak, Blazej
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi. Jagiellonian Univ, Inst Bot, Dept Plant Cytol & Embryol, PL-30387 Krakow, Poland.;Polish Acad Sci, W Szafer Ist Bot, PL-31512 Krakow, Poland..
    Jacobsson, Erik
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Kuta, Elzbieta
    Jagiellonian Univ, Inst Bot, Dept Plant Cytol & Embryol, PL-30387 Krakow, Poland..
    Goransson, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Exogenous plant hormones and cyclotide expression in Viola uliginosa (Violaceae)2015Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 117, s. 527-536Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Plants from Violaceae produce cyclotides, peptides characterized by a circular peptide backbone and a cystine knot. This signature motif gives stability that can harness a wide spectrum of biological activities, with implications in plant defense and with applications in medicine and biotechnology. In the current work, cyclotide expressing in vitro cultures were established from Viola uliginosa. These cultures are useful models for studying biosynthesis of cyclotides and can also be used in their production. The cyclotide expression pattern is shown to be dependent on exogenous plant growth regulators, both on peptide and gene expression levels. The highest yields of cyclotides were obtained on media containing only a cytokinin and were correlated with storage material accumulation. Exposure to auxins decreased cyclotide production and caused shifting of the biosynthesis pattern to root specific cyclotides. The response to stimuli in terms of cyclotide expression pattern appears to be developmental, and related to polar auxin transportation and the auxin/cytokinin ratio regulating tissue differentiation. By the use of whole transcriptome shotgun sequencing (WTSS) and peptidomics, 20 cyclotide sequences from V. uliginosa (including 12 new) and 12 complete precursor proteins could be identified. The most abundant cyclotides were cycloviolacin O3 (CyO3), CyO8 and CyO13. A suspension culture was obtained that grew exponentially with a doubling time of approximately 3 days. After ten days of growth, the culture provided a yield of more than 4 mg CyO13 per gram dry mass.

  • 13.
    Svangård, Erika
    et al.
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Göransson, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Smith, Derek
    Verma, Chandra
    Backlund, Anders
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Bohlin, Lars
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Claeson, Per
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Farmaceutiska fakulteten, Institutionen för läkemedelskemi, Avdelningen för farmakognosi.
    Primary and 3-D modelled structures of two cyclotides from Viola odorata2003Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 64, nr 1, s. 135-142Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    Two polypeptides named vodo M and vodo N, both of 29 amino acids, have been isolated from Viola odorata L. (Violaceae) using ion exchange chromatography and reversed phase HPLC. The sequences were determined by automated Edman degradation, quantitative amino acid analysis, and mass spectrometry (MS). Using MS, it was established that vodo M (cyclo-SWPVCTRNGAPICGESCFTGKCYTVQCSC) and vodo N (cyclo-SWPVCYRNGLPVCGETCTLGKCYTAGCSC) form a head-to-tail cyclic backbone and that six cysteine residues are involved in three disulphide bonds. Their origin, sequences, and cyclic nature suggest that these peptides belong to the family of cyclic plant peptides, called cyclotides. The three-dimensional structures of vodo M and vodo N were modelled by homology, using the experimentally determined structure of the cyclotide kalata B1 as the template. The images of vodo M and vodo N show amphipathic structures with considerable surface hydrophobicity for a protein modelled in a polar environment.

  • 14.
    Söderhäll, Irene
    Uppsala universitet, Teknisk-naturvetenskapliga vetenskapsområdet, Biologiska sektionen, Institutionen för fysiologi och utvecklingsbiologi, Jämförande fysiologi.
    Properties Of Carrot Polyphenoloxidase1995Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 39, nr 1, s. 33-38Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A latent phenoloxidase (PPO) was purified in the presence of protease inhibitors from carrot cells to electrophoretic homogeneity. The inactive enzyme had a M(r) of ca 59 000 under denaturating conditions as judged by SDS-PAGE. When stored in the absence of protease inhibitors, PPO in a crude extract was converted to forms with a lower M(r). Phenol oxidase activity of the purified PPO was induced by the presence in the assay medium of 12.5 mM Tris, and 6 mM CaCl2 increased the activity further.

  • 15. Yoshizaki, Lucila
    et al.
    Troncoso, María F.
    Lopes, Jose L. S.
    Hellman, Ulf
    Uppsala universitet, Medicinska och farmaceutiska vetenskapsområdet, Medicinska och farmaceutiska vetenskapsområdet, centrumbildningar mm, Ludwiginstitutet för cancerforskning.
    Beltramini, Leila M.
    Wolfenstein-Todel, Carlota
    Calliandra selloi Macbride trypsin inhibitor: isolation, characterization, stability, spectroscopic analyses2007Ingår i: Phytochemistry, ISSN 0031-9422, E-ISSN 1873-3700, Vol. 68, nr 21, s. 2625-2634Artikel i tidskrift (Refereegranskat)
    Abstract [en]

    A trypsin inhibitor was purified from Calliandra selloi Macbride seeds (CSTI). SDS-PAGE under non-reducing conditions showed a single band of approximately 20,000Da, while under reducing conditions two bands of 16,000 and 6000Da were observed, indicating that CSTI consists of two polypeptide chains. Molecular masses of 20,078 and 20,279 were obtained by mass spectrometry, although only one pI of 4.0 was observed and one peak was obtained by reversed phase chromatography. Amino-terminal sequence analysis showed homology to Kunitz-type inhibitors. CSTI was able to inhibit trypsin (K(i) 2.21x10(-7)M), alpha-chymotrypsin (K(i) 4.95x10(-7)M) and kallikrein (K(i) 4.20x10(-7)M) but had no effect on elastase. Trypsin inhibitory activity was stable over a wide range of pH and temperature. CSTI was particularly susceptible to DTT treatment, followed by addition of iodoacetamide. Far-UV circular dichroism measurements revealed that CSTI is a beta-II protein. Thermal unfolding showed a two-state transition with a midpoint at 68 degrees C. Far-UV CD spectra of CSTI at pH extremes showed little changes, while more pronounced differences in near-UV CD spectra were detected. Remarkably, treatment with 1mM DTT caused very slight changes in the far-UV CD spectrum, and only after carbamidomethylation was there was a marked loss observed in secondary structure.

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