PubMed Health⌕ Search

Biomedical subjects

E Cederlund

Publications and source records attributed to E Cederlund.

At least 19 recordsLinked to original sources
PubMed1999

Elapid venom toxins: multiple recruitments of ancient scaffolds.

Nigroxins A and B, two myotoxic phospholipases A2 (PLA2s) from the venom of the American elapid Micrurus nigrocinctus, belong to a new PLA2 subclass. Their primary structures were established and compared with those of PLA2s that have already been studied with respect to myotoxic activity. The combination of amino acid residues Arg15, Ala100, Asn108 and a hydrophobic residue at position 109 is present exclusively in class I PLA2s that display myotoxic activity. These residues cluster within a surface region rich in positive charges and are suggested to play a role in the interaction with the target membrane of the muscle fibers. It is concluded that the myotoxic PLA2s resulted from recruitment of an ancient scaffold. Dendrotoxins and alpha-neurotoxins are similarly derived from other old structures, which are, however, now also present in nontoxic proteins that are widely distributed throughout the animal kingdom. The evolutionary pathways by which elapid PLA2s acquired myotoxicity and dendrotoxins acquired K+-channel blocker activity are traced. They demonstrate how existing scaffolds were adapted stepwise to serve toxic functions by exchange of a few surface-exposed residues.

Amino Acid Sequence↗
PubMed1998

Acetyl xylan esterase II from Penicillium purpurogenum is similar to an esterase from Trichoderma reesei but lacks a cellulose binding domain.

Penicillium purpurogenum produces at least two acetyl xylan esterases (AXE I and II). The AXE II cDNA, genomic DNA and mature protein sequences were determined and show that the axe 2 gene contains two introns, that the primary translation product has a signal peptide of 27 residues, and that the mature protein has 207 residues. The sequence is similar to the catalytic domain of AXE I from Trichoderma reesei (67% residue identity) and putative active site residues are conserved, but the Penicillium enzyme lacks the linker and cellulose binding domain, thus explaining why it does not bind cellulose in contrast to the Trichoderma enzyme. These results point to a possible common ancestor gene for the active site domain, while the linker and the binding domain may have been added to the Trichoderma esterase by gene fusion.

Acetylesterase↗
PubMed1997

Characterization of a marsupial glutathione transferase, a class Alpha enzyme from Brown Antechinus (Antechinus stuartii).

The major form of glutathione transferase from the marsupial Antechinus stuartii has been purified and characterized as an Alpha class enzyme (Ast GST A1-1) with distant sequence relationships to other class Alpha sublines, compatible with the early origin of marsupials. Amino acid replacements toward the closest enzyme characterized (chicken, form A3) involve no less than 79 positions (36%). At the active site, as deduced from comparisons with the known tertiary structure of the corresponding human enzyme, over half of the residues (8 of 15) ascribed to substrate binding interactions are exchanged although the general character of that site is conserved, while only 1 of 11 positions ascribed to interactions with GSH is exchanged. Class variability and species variability appear to coincide, with divergent segments centering around positions 33-49, 103-130 and 205-222. The pattern is reminiscent of that in similarly multiple MDR alcohol dehydrogenases. Both these enzyme families involved in cellular defense reactions have diverged considerably.

Amino Acid Sequence↗
PubMed1994

11-Hydroxythromboxane B2 dehydrogenase is identical to cytosolic aldehyde dehydrogenase.

11-Hydroxythromboxane B2 dehydrogenase purified from porcine kidney has been identified as cytosolic aldehyde dehydrogenase (EC 1.2.1.3). This identification is based on protein characteristics, sequence analysis of one proteolytic digest, blocked N-terminus, subunit molecular mass of 55 kDa, and enzymatic activities. The sequence difference with the human enzyme is 7.5% in the fragments analyzed (29 exchanges of 388 positions, corresponding to the expected species variability for cytosolic aldehyde dehydrogenase). The substrate thromboxane B2 contains a hemiacetal in its ring structure, but the reaction most likely proceeds via the aldehyde form of the substrate. This finding is in agreement with the proposed metabolism of 4-hydroxycyclophosphamide and highlights the possibility that molecules containing a hemiacetal structure can function as substrates for aldehyde dehydrogenase.

Alcohol Oxidoreductases↗
PubMed1994

Mammalian class IV alcohol dehydrogenase (stomach alcohol dehydrogenase): structure, origin, and correlation with enzymology.

The structure of a mammalian class IV alcohol dehydrogenase has been determined by peptide analysis of the protein isolated from rat stomach. The structure indicates that the enzyme constitutes a separate alcohol dehydrogenase class, in agreement with the distinct enzymatic properties; the class IV enzyme is somewhat closer to class I (the "classical" liver alcohol dehydrogenase; approximately 68% residue identities) than to the other classes (II, III, and V; approximately 60% residue identities), suggesting that class IV might have originated through duplication of an early vertebrate class I gene. The activity of the class IV protein toward ethanol is even higher than that of the classical liver enzyme. Both Km and kcat values are high, the latter being the highest of any class characterized so far. Structurally, these properties are correlated with replacements at the active site, affecting both substrate and coenzyme binding. In particular, Ala-294 (instead of valine) results in increased space in the middle section of the substrate cleft, Gly-47 (instead of a basic residue) results in decreased charge interactions with the coenzyme pyrophosphate, and Tyr-363 (instead of a basic residue) may also affect coenzyme binding. In combination, these exchanges are compatible with a promotion of the off dissociation and an increased turnover rate. In contrast, residues at the inner part of the substrate cleft are bulky, accounting for low activity toward secondary alcohols and cyclohexanol. Exchanges at positions 259-261 involve minor shifts in glycine residues at a reverse turn in the coenzyme-binding fold. Clearly, class IV is distinct in structure, ethanol turnover, stomach expression, and possible emergence from class I.

Alcohol Dehydrogenase↗
PubMed1992

Class IV alcohol dehydrogenase (the gastric enzyme). Structural analysis of human sigma sigma-ADH reveals class IV to be variable and confirms the presence of a fifth mammalian alcohol dehydrogenase class.

Human gastric alcohol dehydrogenase (sigma sigma-ADH) was submitted to peptide analysis at picomole scale. A total of 72 positions were determined in the protein chain, providing information on three aspects of alcohol dehydrogenase structures in general. First, the data establish the presence of a unique class of the enzyme, now confirmed as class IV, expressed in gastric tissue and separate from another novel class, now termed class V. Second, the class IV gastric enzyme has active site relationships compatible with an ethanol-active, zinc-containing alcohol dehydrogenase. Third, this enzyme class is of the variable type, like that for the 'variable', classical liver alcohol dehydrogenase of class I, and in contrast to that for the 'constant' class III enzyme. Known human alcohol dehydrogenase structures now prove the presence of at least seven human genes for the enzyme and nine for the whole protein family.

Alcohol Dehydrogenase↗
PubMed1991

Characterization of human deoxycytidine kinase. Correlation with cDNA sequences.

Existing data on the structure of human deoxycytidine kinase (dCK) diverge. A monomeric 60 kDa form has been isolated and the cloning of a cDNA coding for 626 amino acids corresponding to a 71 kDa protein has been reported. However, pure dCK isolated from leukemic spleen is a dimer of 30 kDa subunits. Amino acid sequences of peptides from digests of this protein are now presented. None of the peptide structures obtained correspond to the cDNA for the 71 kDa protein, but to a cDNA for a 30.5 kDa dCK recently cloned. Furthermore, homology of the peptide sequences od dCK to parts of thymidine kinases and protein-tyrosine kinases are detected.

Amino Acid Sequence↗
PubMed1991

Amphibian alcohol dehydrogenase, the major frog liver enzyme. Relationships to other forms and assessment of an early gene duplication separating vertebrate class I and class III alcohol dehydrogenases.

Submammalian alcohol dehydrogenase structures can be used to evaluate the origins and functions of the different types of the mammalian enzyme. Two avian forms were recently reported, and we now define the major amphibian alcohol dehydrogenase. The enzyme from the liver of the Green frog Rana perezi was purified, carboxymethylated, and submitted to amino acid sequence determination by peptide analysis of six different digests. The protein has a 375-residue subunit and is a class I alcohol dehydrogenase, bridging the gap toward the original separation of the classes that are observable in the human alcohol dehydrogenase system. In relation to the human class I enzyme, the amphibian protein has residue identities exactly halfway (68%) between those for the corresponding avian enzyme (74%) and the human class III enzyme (62%), suggesting an origin of the alcohol dehydrogenase classes very early in or close to the evolution of the vertebrate line. This conclusion suggests that these enzyme classes are more universal among animals than previously realized and constitutes the first real assessment of the origin of the duplications leading to the alcohol dehydrogenase classes. Functionally, the amphibian enzyme exhibits properties typical for class I but has an unusually low Km for ethanol (0.09 mM) and Ki for pyrazole (0.15 microM) at pH 10.0. This correlates with a strictly hydrophobic substrate pocket and one amino acid difference toward the human class I enzyme at the inner part of the pocket. Coenzyme binding is highly similar, while subunit-interacting residues, as in other alcohol dehydrogenases, exhibit several differences.(ABSTRACT TRUNCATED AT 250 WORDS)

Alcohol Dehydrogenase↗
PubMed1990

Class IV mammalian alcohol dehydrogenase. Structural data of the rat stomach enzyme reveal a new class well separated from those already characterized.

The stomach form of alcohol dehydrogenase has been structurally evaluated by peptide analysis covering six separate regions of the rat enzyme. Overall, this new structure differs widely (32-40% residue differences) from the structures of three classes of alcohol dehydrogenase characterized before from the same species. Consequently, this novel enzyme constitutes a true fourth class of mammalian alcohol dehydrogenase. In particular, differences are extensive also towards class II, although enzymatic and physicochemical properties initially suggested overall similarities with class II. The new structure establishes the presence of one further alcohol dehydrogenase mammalian gene, extends the enzyme family derived from repeated gene duplications, and confirms tissue-specific expressions.

Alcohol Dehydrogenase↗
PubMed1990

Fast atom bombardment mass spectrometry and chemical analysis in determinations of acyl-blocked protein structures.

Peptide generation and fast atom bombardment mass spectrometry in combination with conventional chemical analysis was used to identify the blocking group and establish the N-terminal structure of six different proteins at the nanomole level. In this manner, the first terminal structures of three non-mammalian alcohol dehydrogenases were determined, demonstrating the presence of N-terminal acetylation in these piscine, amphibian, and avian enzymes. Similarly, two different yeast glucose-6-phosphate dehydrogenases and a minor variant of a human alcohol dehydrogenase were found to be acetylated. The exact end location of C-terminal structures was also established. Together, the analyses permit the definition of terminal regions and blocking groups, thus facilitating the delineation of remaining structures.

Acetylation↗
PubMed1989

Cytosolic glutathione transferases from rat liver. Primary structure of class alpha glutathione transferase 8-8 and characterization of low-abundance class Mu glutathione transferases.

Six GSH transferases with neutral/acidic isoelectric points were purified from the cytosol fraction of rat liver. Four transferases are class Mu enzymes related to the previously characterized GSH transferases 3-3, 4-4 and 6-6, as judged by structural and enzymic properties. Two additional GSH transferases are distinguished by high specific activities with 4-hydroxyalk-2-enals, toxic products of lipid peroxidation. The most abundant of these two enzymes, GSH transferase 8-8, a class Alpha enzyme, has earlier been identified in rat lung and kidney. The amino acid sequence of subunit 8 was determined and showed a typical class Alpha GSH transferase structure including an N-acetylated N-terminal methionine residue.

Amino Acid Sequence↗
PubMed1988

Identification of hormone-interacting amino acid residues within the steroid-binding domain of the glucocorticoid receptor in relation to other steroid hormone receptors.

Purified rat liver glucocorticoid receptor was covalently charged with [3H]glucocorticoid by photoaffinity labeling (UV irradiation of [3H]triamcinolone acetonide-glucocorticoid receptor) or affinity labeling (incubation with [3H]dexamethasone mesylate). After labeling, separate samples of the denatured receptor were cleaved with trypsin (directly or after prior succinylation), chymotrypsin, and cyanogen bromide. Labeled residues in the peptides obtained were identified by radiosequence analysis. The peaks of radioactivity corresponded to Met-622 and Cys-754 after photoaffinity labeling with [3H]triamcinolone acetonide and Cys-656 after affinity labeling with [3H]dexamethasone mesylate. The labeled residues are all positioned within hydrophobic segments of the steroid-binding domain. The patterns of hydropathy and secondary structure for the glucocorticoid receptor are highly similar to those for the progestin receptor and similar but less so to those for the estrogen receptor and to those for c-erb A.

Affinity Labels↗
PubMed1988

Primary structure of glycolate oxidase from spinach.

The primary structure of glycolate oxidase from spinach has been determined. Six different types of peptide digest were investigated, utilizing CNBr, proteolytic enzymes, and chemical modifications to change a specificity of cleavage. In total, 90 peptides were purified and analyzed. The studies were aimed at correlation with crystallographic analysis of the same protein carried through in parallel and with cDNA studies which utilized initially determined amino acid sequences for synthesis of oligonucleotide probes. Continuous comparisons with the results from the crystallographic studies helped at an early stage to secure peptide overlaps, at the same time as the peptide data secured residue assignments in the electron density maps. In the end, all data agree and regions from all parts of the molecule have been checked by independent methods of analysis. The primary structure establishes the type of N-terminal post-translational processing, and yields information on segments not fully defined in electron density maps. Combined, the chemical, crystallographic, and cDNA data give extensive reliability. The peptide analysis shows that the N-terminus is blocked by acylation of the initiator methionine, which is in a primary structure typical for non-removal of the methionine in the processing events of the nascent protein chain. The molecule is comparatively rich in menthionine and some other generally less common residues, but has only one cysteine residue and no extensive hydrophobic segment. An amino acid sequence homology with flavocytochrome b2 from yeast, as expected from known similarities in tertiary structure, is observed (33% residue identities).

Alcohol Oxidoreductases↗
PubMed1986

Peptides of postulated inhibin activity. Lack of in vitro inhibin activity of a 94-residue peptide isolated from human seminal plasma, and of a synthetic replicate of its C-terminal 28-residue segment.

A 94-residue polypeptide isolated from human seminal plasma and its chemically synthesized C-terminal 28-residue segment were studied in an in vitro inhibin bioassay utilizing rat pituitary cell cultures. Both peptides have previously been claimed to have inhibin activities, and the effects on the secretion and cellular content of gonadotrophins (FSH and LH) were now assessed in the in vitro assay. No inhibition was found. After 72 h of culture, both the cellular content and the spontaneous as well as the LHRH-stimulated release of bioactive or immunoactive FSH and LH remained unaffected. Similarly, no effects were found on the storage and/or release of prolactin, growth hormone, or thyrotropin. We conclude that both the native 94-residue peptide and the synthetic replicate of its C-terminal 28-residue segment, do not influence the pituitary FSH secretion when assessed in this in vitro system.

Animals↗