Nathan Back

Chromosomal damage in human leukocytes induced by lysergic acid diethylamide.

Addition of lysergic acid diethylamide to cultured human leukocytes resulted in a marked increase of chromosomal abnormalities. The distribution of chromosome breaks deviated significantly from random, with an accumulation of aberrations in chromosome No. 1. Cytogenetic investigation of a patient extensively treated with this drug over a 4-year period for paranoid schizophrenia showed a similar increase in chromosomal damage.

Rodent kinin-forming enzyme systems—I: Purification and characterization of plasma kininogen

Low molecular weight (LMW) kininogen was purified 70-fold with a 16% yield from fresh rat plasma by DEAE-Sephadex chromatography, ammonium sulfate precipitation, Sephadex G-200 gel filtration, SP-Sephadex chromatography, CM-cellulose chromatography, and Sephadex G-200 gel filtration. Ferguson plots of polyacrylamide gel electrophoretic patterns revealed four bands with relative molecular weights of 64,000, 123,500, 252,436 and 357,900 (ratio of 1:2:4:6). Sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis provided a single protein band with a molecular weight of 72,000, suggesting that the four kininogen bands had been caused by the aggregation of a single oligomeric protein. The purified LMW rat kininogen Fraction B (3.9 micrograms bradykinin/mg) was used to elicit an antiserum in the rabbit. Monospecificity of the antiserum was demonstrated by immunoelectrophoresis (Laurell rocket and Grabar methods) and, thus, the homogeneity of the kininogen was also. The purified kininogen (both Fractions A and B) formed kinin with human urinary kallikrein, rat urinary kallikrein and hog pancreatic kallikrein. Murphy-Sturm lymphosarcoma acid protease also formed kinin when incubated with the kininogen at pH 3.0. The isoelectric point for both fractions was at pH 4.3. Amino acid analyses showed the two kininogen fractions to be rich in acidic amino acids and to have a total carbohydrate content of 8.5% consisting of galactose (1.2 to 1.5%), mannose (1.9 to 2.1%), N-acetylglucosamine (4.3 to 5.1%), N-acetylgalactosamine (0.3%), and sialic acid (0.68%).

Effect of apotinin, EACA and heparin on growth and vasopeptide system of murphy—sturm lymphosarcoma☆☆☆

The effect of aprotinin, EACA and heparin on the growth of the transplanted rodent Murphy-Sturm lymphosarcoma and on several components of the tumor kinin-forming enzyme system was studied. Therapy was administered for 15 days, 3 times daily, one day after tumor transplant. Tumor weights and biochemical parameters (prekallikrein, kininogen and kininase) were measured on the 8th, 10th, 12th and 15th day of therapy. Both aprotinin and heparin significantly inhibited (p less than 0.01) tumor growth by the 15th day, compared to control, saline-administered animals. EACA did not affect the tumor growth rate at any time period. Tumor prekallikrein and kininogen levels were significantly higher (p less than 0.01) in the aprotinin-treated animals. Kininogen levels were higher in the EACA-treated tumors during the latter phase of growth. None of the agents affected the increase in tumor kininase activity that occured during the growth of the tumors. The tumor inhibition and biochemical data suggest the involvement of proteases in the neoplastic process.

Genome instability in cancer development

Part 1. The Problem of Genome Instability 1.1. The Multiplicity of Mutations in Human Cancers by Ranga N. Venkatesan and Lawrence A. Loeb 1.2. Monitoring chromosome rearrangements by Michael R. Speicher Part 2. DNA Repair and Mutagenesis 2.1. Nucleotide excision repair and its connection with cancer and ageing by Jaan-Olle Andressoo, Jan H.J. Hoeijmakers and Harm de Waard 2.2. DNA mismatch repair and colon cancer by Giancarlo Marra and Josef Jiricny 2.3. Base excision repair by Lisiane B. Meira, Nicholas E. Burgis and Leona D. Samson 2.4. DNA double strand break repair by Penny A. Jeggo 2.5. Translesion synthesis and error-prone polymerases by Catherine M. Green and Alan R. Lehmann Part 3. Cell Cycle Progression and Chromosome Aberrations 3.1. The INK4A/ARF network - cell cycle checkpoint or emergency brake? By Ana Gutierrez del Arroyo and Gordon Peters 3.2. DNA replication and genomic instability by Wenge Zhu, Tarek Abbas and Anindya Dutta 3.3. The dream of every chromosome: equal segregation for a healthy life of the host by Tomohiro Matsumoto and Mitsuhiro Yanagida 3.4. Telomere structural dynamics in genome integrity control and carcinogenesis by Roger A. Greenberg and K. Lenhard Rudolph 3.5. Gene amplification mechanisms by Michelle Debatisse and Bernard Malfoy 3.6. DNA methylation and cancer-associated genetic instability by Melanie Ehrlich 3.7. Deregulation of the centrosome cycle and the origin of chromosomal instability in cancer by Wilma L. Lingle, Kara Lukaswiewicz and Jeffrey L. Salisbury Part 4. Genome Integrity Checkpoints 4.1. Mammalian DNA damage response pathway by Zhenkun Lou and Junjie Chen 4.2. ATM and cellular response to DNA damage by Martin F Lavin, Sergei Kozlov, Nuri Gueven, Cheng Peng, Geoff Birrell, Phillip Chen and Shaun Scott 4.3 Kinetochore function, chromosome segregation and the spindle assembly checkpoint by Tim J. Yen and Gary D. Kao

Dipeptidyl Aminopeptidases in Health and Disease

Structure and Function of Dipeptidyl Aminopeptidases. Dipeptidyl Peptidase IV Substrates I. de Meester, et al. Structure-Function Relationship of DPP IV: Insights into its Dimerisation and Gelatinase Activity O. Baum, et al. Exploration of the Active Site of Dipeptidyl Peptidase IV From Porphyromonas gingivalis A.-M. Lambeir, et al. Modification of the Biological Activity of Chemokines by Dipeptidyl Peptidase IV - a Side Effect in the Use of Inhibitors? R. Mentlein, et al. Molecular Chimeras and Mutational Analysis in the Prolyl Oligopeptidase Gene Family K. Ajami, et al. The Specificity of DP IV for Natural Substrates is Peptide Structure Determined K. Kuhn-Wache, et al. New Results on the Conformations of Potent DP IV (CD26) Inhibitors Bearing the N-terminal MWP Structural Motif C. Mrestani-Klaus, et al. Different Inhibition Mechanisms of Dipeptidyl Peptidase IV by Tryptophan Containing Peptides and Amides A. Stockel-Maschek, et al. Re-Uptake Mechanisms of Peptide Fragments after DPP IV-Mediated Proteolysis in the Peripheral Nervous System Q. Thai Dinh, et al. DPP IV-Like Enzymes. Dipeptidyl Peptidase IV Gene Family Tong Chen, et al. Seprase-DPPIV Association and Prolyl Peptidase and Gelatinase Activities of the Protease Complex G. Ghersi, et al. Dipeptidyl Peptidase-IV Activity and/or Structure Homologues (DASH) in Transformed Neuroectodermal Cells R. Malik, et al. Characterisation of Human DP IV Produced by a Pichia pastoris Expression System J.W. Baer, et al. Isolation and Characterization of Attractin-2 D. Friedrich, et al. Investigation of DP IV-dependent Protein-Protein Interactions using Surface Plasmon Resonance J. Stork, et al. Immune Mechanisms andImmune Disorders. Synergistic Action of DPIV and APN in the Regulation of T Cell Function U. Lendeckel, et al. CD26/DPP IV in Experimental and Clinical Organ Transplantation S. Korom, et al. CD26 is Involved in the Regulation of T-Cell Plasma Membrane Compartmentation J. Lojo, et al. Inhibition of Dipeptidylpeptidase IV (DPP IV, CD26) Activity Modulates Surface Expression of CTLA-4 in Stress-Induced Abortions J. Ruter, et al. Dipeptidyl Peptidase IV/CD26 in T Cell Activation, Cytokine Secretion and Immunoglobulin Production Hua Fan, et al. Dipeptidyl Peptidase IV Inhibitors with the N-terminal MXP Sequence: Structure-Activity-Relationships J. Faust, et al. On the Role of Dipeptidyl Peptidase IV in the Digestion of an Immunodominant Epitope in Celiac Disease S. Koch, et al. The Properties of Human and Bovine CD8+CD26+ T Cells Induced by a Microbial Superantigen Sang-Un Lee, et al. Angiogenesis and Cancer. DPPIV and Seprase in Cancer Invasion and Angiogenesis W.-T. Chen. Glutamate Carboxypeptidase II Inhibition as a Novel Therapeutic Target C. Rojas, et al. Dual Role of Dipeptidyl Peptidase IV (DPP IV) in Angiogenesis and Vascular Remodeling J. Kitlinska, et al. CD26 Expression on Cutaneous Infiltrates from Patients with Cutaneous T-Cell Lymphoma (CTCL) M. Novelli, et al. Intrahepatic Expression of Collagen and Fibroblast Activation Protein (FAP) in Hepatitis C Virus Infection M.D. Gorrell, et al. Expression of CD26/Dipeptidyl Peptidase IV in Endometrial Adenocarcinoma and its Negative Correlation with Tumor Grade H. Kajiyama, et al. Adhesion Potency to Mesothelial Cells by Overexpression of Dipeptidyl Peptidase IV F. Kikkawa, et al. Survival Time and Invasive Activity due to Dipeptidyl Peptidase IV Overexpression in Ovarian C

Rodent kinin-forming enzyme systems--II. Purification and characterization of an acid protease from Murphy-Sturm lymphosarcoma.

An acid protease from the rat Murphy-Sturm lymphosarcoma (MSLS) tumor was purified 640-fold by extraction of the tumor tissue, acid precipitation with glacial acetic acid, ammonium sulfate precipitation, DEAE-Sephadex A-50 batch adsorption, QAE-Sephadex A-50 column chromatography, Sephadex G-200 gel filtration, and CM-32 cellulose chromatography. The protease hydrolyzed bovine hemoglobin and formed vasopeptide kinins when incubated with purified rat plasma kininogen. Two protease fractions obtained by Sephadex G-200 gel filtration had identical molecular weights of 39,500-41,000 and were similar in other physico-chemical and kinetic characteristics. The purified enzyme showed three major isozymic forms (alpha, beta and gamma) with isoelectric points (pI) of 5.2, 5.5 and 5.8, respectively, and nearly identical amino acid compositions. The enzyme had a high moles percent of both aspartic and glutamic acids. The carbohydrate moiety of the enzyme contained 2 moles of N-acetylglucosamine and 8 moles of mannose per mole of enzyme. The pH optimum for the digestion of bovine hemoglobin was approximately 3.0 with a sharp decline of activity on either side of the pH optimum. The protease activity was very stable above pH 3.4. The Km values for the purified enzyme fractions A and B were 31.17 and 31.19 microM, respectively, and the corresponding Vmax values were 6.17 and 5.5 microM tyrosine per mg per min at 37 degrees and pH 3.0. The enzyme was inhibited strongly by pepstatin (Ki = 31 X 10(-9)M and alpha = 0.1). The acid protease released kinin from purified rat plasma kininogen at an initial rapid rate which plateaued at 460 ng bradykinin equivalents/mg substrate after a 2-hr incubation at 37 degrees.

Fluorescent N-methylanthranilyl (Mantyl) tag for peptides : its application in subpicomole determination of kinins

Highly fluorescent N-methylanthranilyl (Mantyl) peptide derivatives were prepared by a one-step reaction with N-methylisatoic anhydride (MIA) for quantitative detection in HPLC. Reactions were carried out in an organic medium of acetonitrile-triethylamine, in aqueous alkaline sodium carbonate and sodium phosphate buffers. 4-Dimethylaminopyridine (DMAP) catalyzed specific mantylation of -NH2 groups of peptides in the organic reaction medium. The DMAP had no effect in the aqueous buffered reaction systems. Proline amino-terminal peptides reacted equally well with MIA. Mantyl-bradykinin had excitation and fluorescence maxima at 350 nm and 426 nm in water and water/acetonitrile (ACN)/trifluoroacetic acid (TFA) solvent mixtures, respectively. Fluorescence intensity increased with an increase in ACN concentration and decreased with an increase in acid content. Mantyl kinins were completely resolved on a C18 reversed phase HPLC column using an ACN-0.1% TFA gradient and their behavior on the column was similar to having an extra amino acid. Di-Mantyl derivatives obtained with Lys-BK and Met-Lys-BK did not exhibit fluorescence appreciably higher than Mantyl-BK. Fluorescence detection of Mantyl kinins was about 50-100 times more sensitive (lower limits of 0.1 to 0.5 picomole) than UV detection of the phenylisothiocyanate-derivatized kinins under typical HPLC conditions.

The Vasopeptide Kinin System in Acute Clinical Cardiac Diseases

Most deaths in patients with acute myocardial infarction (AMI) have been due to severe arrhythmia and cardiogenic shock. While continuous electrocardiographic monitoring and drug treatment have significantly reduced mortality due to arrhythmia, cardiogenic shock still remains a major cause of death in patients with AMI.(1).

The Influence of Blood pH on Peripheral Vascular Tone: Possible Role of Proteases and Vaso-Active Polypeptides

The ability of the microvasculature to respond to the ever changing metabolic needs of the various tissues is a fundamental requirement for maintaining homeostasis. The underlying mechanism is probably a complex one, involving a multiplicity of mediators, the release of which may be initiated by a primary metabolic stimulus.

Purification and characterization of kinin-forming acid protease from mouse fibroblasts L-929

Abstract Purification and further characterization was carried out on a kinin-forming acid protease isolated from a rodent fibroblast cell line L-929 grown in stationary cell culture (N. Back and R. Steger,[7]). The cells, cultured in minimal essential medium containing 10% fetal calf serum and 0.4% lactalbumin, were homogenized, the homogenate dialyzed for 18 hr against 0.01 M phosphate buffer at pH 6.8 in 0.1 M NaCl and 1.0 mM EDTA, and centrifuged at 10000 rpm for 45 min. The supernatant, which digested denatured hemoglobin at pH 4.0, was fractionated first on a G-200 Sephadex column. Kinin-forming activity, compared with that of the supernatant on an isolated perfused rat uterus preparation, was identified in fractions 25–40 when incubated for 24 hr at pH 4.0 with rat plasma kininogen substrate. The active fractions were pooled and purified further on a hydroxy-patite column. Treatment of the active fractions with 5 mM cysteine increased the activity 2-fold. Final purification was carried out on a DEAE-A50 Sephadex ion exchange column. The purification factor, compared to the initial supernatant, was 9.4 with a 13.8% yield and a specific activity of 2062.5 ng kinin per mg protein. Dialyzed and centrifuged rat plasma fractionated on a DEAE-A50 Sephadex column initially yielded two apparent kininogen species which resolved into a single major molecular species following passage through a G-100 Sephadex column. The purified enzyme and substrate preparations were used to establish the optimum kinin-forming activity at pH 3.8–4.0. The molecular weights of the enzyme and kininogen were estimated on a G-200 Sephadex column to be 38000–39000 and 115000 respectively. The amount of kinin formed was a function of incubation time and enzyme concentration. The acid protease activity was found localized primarily in the 10000 g supernatant cell fraction. The 500 g cell fraction also exhibited activity.