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H F Deutsch

Publications and source records attributed to H F Deutsch.

16 recordsLinked to original sources
PubMed1991

Preparation of human manganese superoxide dismutase by tri-phase partitioning and preliminary crystallographic data.

Human liver manganese superoxide dismutase has been purified by a short procedure that includes a tri-phase partitioning step to provide materials that can be crystallized from ammonium sulfate. X-ray diffraction studies at 3 A resolution show that the crystals belong to the hexagonal space group P6(1)22 or P6(5)22, with cell dimensions a = b = 81.1 A, c = 242.2 A. Manganese superoxide dismutase levels as determined by enzymatic assay as well as by enzyme-linked immunosorbent assay indicated that considerable variations occur in different livers but the total superoxide dismutase activity (Mn superoxide dismutase plus Cu,Zn superoxide dismutase) seems to be kept at constant values.

Ammonium Sulfate↗
PubMed1991

Analyses of polypeptides in the liver of a novel mutant (LEC rats) to hereditary hepatitis and hepatoma by two-dimensional gel electrophoresis: identification of P29/6.8 as carbonic anhydrase III and triosephosphate isomerase.

1. Total cellular proteins from the livers of 4-, 16- and 52-week-old hepatitis- and hepatoma-predisposed Long-Evans Cinnamon (LEC) rats were compared to those from the livers of the corresponding control rats [Long-Evans Agouti (LEA) rats] by two-dimensional gel electrophoresis. 2. A polypeptide, p50/7.2 (molecular weight x 10(-3)/isoelectric point) was only found in the LEC rats, and the p43/6.4 component was greater and the p51/6.8 component was less in the LEC rats than in the LEA rats during aging. 3. A polypeptide, p29/6.8, was dramatically greater in 4-week-old LEC rats than in 4-week-old LEA rats. 4. By sequencing and Western blotting analysis, the marked differences in the level of the p29/6.8 component were found to be due to carbonic anhydrase III.

Aging↗
PubMed1991

Chemistry and biology of alpha-fetoprotein.

Alpha-Fetoprotein (AFP) is a product of specific fetal tissues and of neoplastic cells of hepatocyte or germ cell origin in adults. This protein belongs to a gene family that is phylogenetically most closely related to serum albumin. Its primary, secondary, and tertiary structural aspects appear similar to the three-domain concept proposed for the latter protein. The primary sequence of AFP departs most widely from serum albumin in the first 135 amino acid residues, with about 42% of the remaining 590 residues of the human proteins being identical. Some evidence exists that there are limited sequence differences in the AFP of a given animal species. AFP shows considerable charge heterogeneity that appears to relate mostly to its glycoid moiety. The proteins of some species such as the rat show more pronounced heterogeneities than that of humans. The variations in extent and type of glycosylations are evidenced by differences in the binding to various lectins. These interactions are being extensively explored in attempts to differentiate the sources of the protein produced by various normal and neoplastic cells and may provide valuable diagnostic methods. AFP, like serum albumin, shows relatively strong binding affinities for a variety of ligands. The most notable difference is the strong preferential binding of polyunsaturated fatty acids by AFP. This protein may play a role in transporting these substances to developing and to malignant cells. Various agents affect the synthesis of this protein both by specific fetal tissues and by neoplastic cells. Marked differences in the responses of cells, particularly those of neoplastic types, are indicative of variations in the genetic factors responsible for control of its synthesis. The subject of the genomic repression of the synthesis of AFP seen in fetal life upon maturation of the liver and the reoccurrence of synthesis upon malignant conversion of hepatocytes and of certain germ cells are of particular interest. The regulation of the closely related AFP and albumin genes is providing a powerful and attractive model to examine molecular events in the activation and inactivation of specific genes during development and in oncogenic processes. Extensive measurements of AFP during pregnancy and in the course of neoplasias, notably hepatoma, are being made to aid in following changes in such developments. Various specific physiological roles for this protein are also being proposed. One of these is its possible action in the regulation of immune processes.(ABSTRACT TRUNCATED AT 400 WORDS)

Amino Acid Sequence↗
PubMed1990

Human liver manganese superoxide dismutase. Purification and crystallization, subunit association and sulfhydryl reactivity.

Manganese superoxide dismutase (Mn-SOD) has been purified with a high yield (320 mg) from human liver (2 kg) and crystallized. Low-angle laser light scattering of the enzyme has shown that native enzyme is a tetrametic form. Four of the eight cysteine residues in the tetramer reacted with 5,5'-dithiobis(2-nitrobenzoic acid) or with iodoacetamide. The others were only reactive in protein heated with SDS or urea after reduction with dithiothreitol or 2-mercaptoethanol. The reactive sulfhydryl group was found to be located at Cys196 by amino acid sequence analysis of Nbs2-reactive peptides isolated by activated thiol-Sepharose covalent chromatography. Incubation of Mn-SOD in 1% SDS for 2 or 3 days at 25 degrees C or 5 min at 100 degrees C gave material showing two prominent components on polyacrylamide gel electrophoresis in the presence of 0.1% SDS. The major component had a molecular mass of 23 kDa; the other, 25 kDa. Reduction of the protein by dithiothreitol or 2-mercaptoethanol heated in SDS produced only the 25-kDa monomer species. Essentially, no thiol groups were detected in the 23-kDa form, in which two cysteine residues appear to have been oxidized to form an intrasubunit disulfide. This indicates that Cys196 has a reactive sulfhydryl and appears to be a likely candidate for a mixed disulfide formation in vivo.

Chromatography, Gel↗
PubMed1990

Purification of soluble enzymes from erythrocyte hemolysates by three phase partitioning.

1. The three phase partitioning method of protein fractionation was successfully applied to human erythrocyte hemolysates for the removal of hemoglobin and the concentration of soluble enzymes. 2. Human carbonic anhydrase I and II, catalase and superoxide dismutase were recovered free of hemoglobin and in good yield in the initial partitioning step, with a 60- to 80-fold enrichment of enzyme activities. 3. After further purification, carbonic anhydrases I and II were obtained at overall yields of 84 and 29%, respectively, crystallized catalase at 38% and superoxide dismutase at 52%.

Carbonic Anhydrases↗
PubMed1979

Human alpha-fetoprotein. Fluorescence studies on binding and proximity relationships for fatty acids and bilirubin.

The binding of bilirubin and the polyene fatty acids cis-parinaric acid and cis-eleostearic acid to human alpha-fetoprotein was studied using fluorescence quenching and fluorescence enhancement techniques. alpha-Fetoprotein has three fatty acid binding sites of decreasing affinity (association constants 2.1 x 10(7) M-1 9.1 X 10(5) M-1, and 1.4 x 10(5) M-1) and one relatively strong and one relatively weak bilirubin binding site (association constants 1.1 x 10(7) M-1 and 1.8 x 10(5) M-1). These association constants are slightly weaker than the corresponding association constants for binding to human albumin. Competition experiments failed to show preferential binding of polyunsaturated fatty acids. Fluorescence quenching was used to determine 11 ligand-ligand and ligand-tryptophanyl residue distances. Each of these 11 calculated distances (ranging from 19 A to 32 A) was within 5 A of the corresponding distances measured previously for human albumin (Berde, C.B., Hudson, B.S., Simoni, R.D., and Sklar, L.A. 1979, J. Biol. Chem. 254, 391-400). Thus, in addition to previously described sequence homology, immunologic cross-reactivity, and other similarities, human albumin and human alpha-fetoprotein have functional and geometric homologies.

Bilirubin↗
PubMed1978

The presence of fatty acids in human alpha-fetoprotein.

alpha-Fetoprotein has been prepared from human fetal tissue by procedures utilizing DEAE-Sephadex, concanavalin A-Sepharose, and isoelectric focusing. A major and a minor component with isoelectric points of 4.7 and 5.3, respectively, have been isolated and are similar to those prepared under various conditions by other investigators. The 4.7 material contains 2.4 mol of fatty acids/mol of protein, whereas the minor component is fat-free. The relative amounts of fatty acid vary somewhat with different preparations. The ranges found in three isolates were as follows: palmitic acid (8 to 11%), stearic acid (2 to 5%), oleic acid (10 to 28%), linoleic acid (7 to 15%), arachidonic acid (12 to 39%), and 4,7,10,13,16,19-docosahexaenoic acid (16 to 42%). Human fetal serum albumin contained 0.7 mol of fatty acid/mol of protein, with arachidonic acid and the docosahexaenoic acid comprising only 11.4% of the total. Removal of fatty acids by treatment with charcoal converted alpha-fetoprotein into material with an isoelectric point of pH 5.3. Addition of arachidonic acid to the lipid-free protein restored it to protein with a pH 4.7 isoelectric point, typical of the major native component. The possible role of the fatty acids in alpha-fetoprotein on the inhibition of various lymphocyte functions is projected.

Chromatography, Gas↗
PubMed1977

Purification and properties of a polymorphic high activity equine erythrocyte carbonic anhydrase.

A polymorphic form of the high activity or C-type of horse erythrocyte carbonic anhydrase has been isolated. It has been designated C2 and differs from the usual C1 form by having a cysteine replacement for arginine at residue 180. This second cysteine, unlike the other, is highly reactive. Isolation of the C2 isozyme by the usual methods results in most of it forming a mixed disulfide with glutathione and this product designated as C3 has an increased anodic mobility. The enzymatic activity and immunologic reactivity of both the C2 and C3 components are the same as for the usual C1 form of the enzyme. The C2 form can be stabilized by alkylation and the carboxamidomethyl derivative has been isolated in crystalline form.

Amino Acid Sequence↗
PubMed1976

Conformational isomerism, rotational allomerism, and divergent evolution in immunoglobulin light chains.

Immunoglobulin light chains are examples of single polypeptide chains synthesized under the control of two genes. The three-dimensional structure of a human (Mcg) lambda-type chain (Bence-Jones) dimer supports the hypothesis of a common primordial gene for the amino ("variable" or V) and carboxyl ("constant" or C) halves of each monomer. However, sequence homologies have been obscurred by divergent evolution of the V and C regions ("domains"). The types of evolutionary changes that have occurred in the domain can be surmised by a comparison of the sequences, using the three-dimensional structures as a basis for alignment. Despite substantial differences in sequences, the hydrophobic character of key internal sites has been maintained in each domain. Regions present in only one domain are situated in position appropriate for their functions, but not deleterious to the general structural integrity of a common fold. The divergence of the V and C domains can be interpreted in terms of rotational allomerism. The cylinders of beta-pleated sheets have rotated in such a way that homologous regions in the two domains perform different functions in their interactions with a second molecule of light or heavy chain. These regions include complementarity-determining sites for antigen binding in the V domains and crossover sites stabilizing dimer formation in the C domains. Differences in surface properties between the V1-V2 and C1-C2 dimeric modules may partially explain why the V regions have been implicated in the formation of amyloid fibrils and in the characteristic thermal behavior of Bence-Jones proteins.

Amino Acid Sequence↗
PubMed1975

Carbonic anhydrase isozymes in American ponies and riding horses: a new polymorphic high-activity type isozyme.

A study of the erythrocyte carbonic anhydrases of 219 American ponies and 76 riding horses has revealed the presence of five variants of the low-activity CA B isozyme and two variants of the high-activity CA C isozyme. The previously undetected variant of CA C was found only in the pony population and had an allele frequency of 8.9%. A family study of animals possessing the CA B variant A2 showed an unexpected high frequency of inheritance.

Alleles↗
PubMedDate not supplied

A three-step purification of manganese superoxide dismutase from human liver on both large and small scales.

A new method for the purification of manganese superoxide dismutase from human liver is described. The procedure involves essentially three steps: DEAE-cellulose, hydroxylapatite, and butyl-Toyopearl chromatographies. The method has several advantages: (i) its simplicity and rapidity (it takes less than 3 days), (ii) its high yield (62%) with a high specific activity (5660 units/mg of purified SOD), and (iii) its suitability for both large- and small-scale purifications.

Chromatography↗