1. |
- Takahashi, Masayuki, 1949, et al.
(author)
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COORDINATION OF MULTIPLE DNA-MOLECULES IN RECA FIBER EVIDENCED BY LINEAR DICHROISM SPECTROSCOPY
- 1991
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In: Biochimie. - : Elsevier BV. - 0300-9084. ; 73:2-3, s. 219-226
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Journal article (peer-reviewed)abstract
- Polarized light spectroscopy has been used to study the interaction of RecA protein with DNA. Several different DNA complexes have been identified and characterized with respect to stoichiometries, base orientation and nuclease accessibility. By using spectroscopically distinguishable DNAs, we determined the number of DNA molecules co-ordinated by the RecA fiber in each of these complexes, and established their base pairing abilities. Based on these observations, we discuss the molecular mechanism of the RecA-mediated strand exchange reaction.
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2. |
- Abdulkarim, Farhad, et al.
(author)
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Missense substitutions lethal to essential functions of EF-Tu
- 1991
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In: Biochimie. - : Elsevier BV. - 0300-9084 .- 1638-6183. ; 73:12, s. 1457-1464
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Journal article (peer-reviewed)abstract
- We have used a simple selection and screening method to isolate function defective mutants of EF-Tu. From 28 mutants tested, 12 different missense substitutions, individually lethal to some essential function of EF-Tu, were identified by sequencing. In addition we found a new non-lethal missense mutation. The frequency of isolation of unique mutations suggests that this method can be used to easily isolate many more. The lethal mutations occur in all three structural domains of EF-Tu, but most are in domain II. We aim to use these mutants to define functional domains on EF-Tu.
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3. |
- Andersson, Rolf E, et al.
(author)
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Antibacterial activity of plantaricin SIK-83, a bacteriocin produced by Lactobacillus plantarum
- 1988
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In: Biochimie. - : Elsevier BV. - 0300-9084 .- 1638-6183. ; 70:3, s. 381-390
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Journal article (peer-reviewed)abstract
- Lactobacillus plantarum SIK-83 produces a bacteriocin, designated plantaricin SIK-83, which inhibits 66 of 68 lactic acid bacteria from the genera Lactobacillus, Leuconostoc, Pediococcus and Streptococcus. A 500-fold dilution of L. plantarum SIK-83 MRS culture supernatant with phosphate buffer was sufficient to kill 105 cells/ml of Pediococcus pentosaceus within 120s. The killing of a sensitive population followed exponential kinetics. It shown that the bacteriocin binds specifically to sensitive cells but not to nonsensitive lactic acid bacteria, the producer strain or Gram-negative bacteria. Sensitive cells, after exposure to the bacteriocin, could be rescued by treatment with proteolytic enzymes. In buffer, plantaricin SIK-83 was adsorbed to the cell surface almost immediately, and morphological lesions were observed within 2 h after the cells were exposed to the bacteriocin. The lethal mode of action appeared to be due to damage to the cell membrane, resulting in cell lysis, which was detected by electron microscopy and by determination of released intracellular components. © 1988.
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4. |
- Bilgin, Neş'e, et al.
(author)
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Mutations in ribosomal proteins L7/L12 perturb EF-G and EF-Tu functons
- 1988
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In: Biochimie. - : Elsevier BV. - 0300-9084 .- 1638-6183. ; 70:5, s. 611-618
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Journal article (peer-reviewed)abstract
- In vitro cycling rates of E. coli ribosomes and of elongation factors EF-Tu and EF-G have been obtained and these are compatible with translation rates in vivo. We show that the rate of translocation is faster than 50 s-1 and therefore that the EF-G function is not a rate limiting step in protein synthesis. The in vivo phenotype of some L7/L12 mutants could be accounted for by perturbed EF-Tu as well as EF-G functions. The S12 mutants that we studied were, in contrast, only perturbed in their EF-Tu function, while their EF-G interaction was not impaired in relation to wild type ribosomes.
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6. |
- Dahlgren, Ann, et al.
(author)
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A novel mutation in ribosomal protein S4 thataffects the function of a mutated RF1
- 2000
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In: Biochimie. - : Société de chimie biologique. - 0300-9084. ; 82:8, s. 683-691
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Journal article (peer-reviewed)abstract
- Release factors (RF) 1 and 2 trigger the hydrolysis of the peptide from the peptidyl-tRNA during translation termination. RF1 binds to the ribosome in response to the stop codons UAG and UAA, whereas RF2 recognizes UAA and UGA. RF1 and RF2 have been shown to bind to several ribosomal proteins. To study this interaction in vivo, prfA1, a mutant form of RF1 has been used. A strain with the prfA1 mutation is temperature sensitive (Ts) for growth at 42 °C and shows an increased misreading of UAG and UAA. In this work we show that a point mutation in ribosomal protein S4 can, on the one hand, make the RF1 mutant strain Ts+; on the other hand, this mutation increases the misreading of UAG, but not UAA, caused by prfA1. The S4 mutant allele, rpsD101, is a missense mutation (Tyr51 to Asp), which makes the cell cold sensitive. The behaviour of rpsD101 was compared to the well-studied S4 alleles rpsD12, rpsD14, and rpsD16. These three mutations all confer both a Ts (44 °C) phenotype and show a ribosomal ambiguity phenotype, which rpsD101 does not. The three alleles were sequenced and shown to be truncations of the S4 protein. None of the three mutations could compensate for the Ts phenotype caused by the prfA1 mutation. Hence, rpsD101 differs in all studied characteristics from the three above mentioned S4 mutants. Because rpsD101 can compensate for the Ts phenotype caused by prfA1 but enhances the misreading of UAG and not UAA, we suggest that S4 influences the interaction of RF1 with the decoding center of the ribosome and that the Ts phenotype is not a consequence of increased readthrough.
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9. |
- Karlsson, E., et al.
(author)
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Snake toxins with high selectivity for subtypes of muscarinic acetylcholine receptors
- 2000
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In: Biochimie. - 0300-9084 .- 1638-6183. ; 82:9-10, s. 793-806
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Journal article (peer-reviewed)abstract
- There are five subtypes of muscarinic acetylcholine receptors (M-1 to M-5) which control a large number of physiological processes, such as the function of heart and smooth muscles, glandular secretion, release of neurotransmitters, gene expression and cognitive functions as learning and memory. A selective ligand is very useful for studying the function of a subtype in presence of other subtypes, which is the most common situation, since a cell or an organ usually has several subtypes. There are many non-selective muscarinic ligands, but only few selective ones. Mambas, African snakes of genus Dendroaspis have toxins, muscarinic toxins, that are selective for M-1, M-2 and M-4 receptors. They consist of 63-66 amino acids and four disulfides which form four loops. They are members of a large group of snake toxins, three-finger toxins; three loops are extended like the middle fingers of a hand and the disulfides and the shortest loop are in the palm of the hand. Some of the toxins target the allosteric site which is located in a cleft of the receptor molecule close to its extracellular part. A possible explanation to the good selectivity is that the toxins bind to the allosteric site, but because of their size they probably also bind to extracellular parts of the receptors which are rather different in the various subtypes. Some other allosteric ligands also have good selectivity, the alkaloid brucine and derivatives are selective for M-1, M-3 and M-4 receptors. Muscarinic toxins have been used in several types of experiments. For instance radioactively labeled M-1 and M-4 selective toxins were used in autoradiography of hippocampus from Alzheimer patients. One significant change in the receptor content was detected in one region of the hippocampus, dentate gyrus, where M-4 receptors were reduced by 50% in patients as compared to age-matched controls. Hippocampus is essential for memory consolidation. M-4 receptors in dentate gyrus may play a role, since they decreased in Alzheimers disease which destroys the memory. Another indication of the role of M-4 receptors for memory is that injection of the M-4 selective antagonist muscarinic toxin 3 (M-4-toxin 1) into rat hippocampus produced amnesia.
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10. |
- Spillmann, Dorothe
(author)
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Heparan sulfate : anchor for viral intruders?
- 2001
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In: Biochimie. - 0300-9084 .- 1638-6183. ; 83:8, s. 811-817
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Journal article (peer-reviewed)abstract
- Heparan sulfates (HS) are ubiquitous, polyanionic carbohydrate chains linked to core proteins in cell membranes and extracellular matrices of all eukaryotes. Due to the complex nature of the HS-biosynthesis, a wealth of different structures are produced. These seem to have a well defined distribution in different tissues and cells throughout development. Binding of endogenous proteins with different functional properties such as growth factors, adhesion molecules or enzymes, is one of the functions of HS. Besides interaction with endogenous factors, glycosaminoglycans (GAG) and especially HS have also been demonstrated to function as receptors for a number of different pathogens. What roles may HS play in the pathogenesis and tropism of different intruders like parasites or viruses? What implications does binding of viruses to HS have for the development of drugs or the application of viral vectors for gene targeting? In this review an attempt is made to collect our present knowledge on viral usage of HS and the implications that follow.
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17. |
- Baldassarre, Maurizio, et al.
(author)
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Amyloid fibril formation by bovine alpha(1)-acid glycoprotein in a reducing environment : The role of disulfide bridges on the observed aggregation kinetics
- 2015
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In: Biochimie. - : Elsevier BV. - 0300-9084 .- 1638-6183. ; 118, s. 244-252
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Journal article (peer-reviewed)abstract
- Bovine alpha(1)-acid glycoprotein (bAGP), a thermostable counterpart of its human homologue, is a positive acute phase protein involved in binding and transportation of a large number of bin-active molecules and drugs across the body. We have investigated the effect of low pH and reducing conditions on the structure of the protein and found that it aggregates at high temperatures. The aggregates show a fibrillar structure when observed with electron microscopy. Aggregation assays using the amyloid-specific dye Thioflavin T show the presence of a lag phase which was neither abolished nor shortened when seeds were added. A priori reduction of the two disulfide bridges of bAGP, on the other hand, abolished the lag phase and reveals a connection between the kinetics of reduction and aggregation. We provide a kinetic interpretation and the corresponding rate laws allowing to model the process of fibril formation by bAGP under reducing conditions. Our interpretation allows to assess the role of disulfide bridges on the fibrillation kinetics of bAGP and can provide a more accurate interpretation of the fibrillation kinetics of other amyloidogenic proteins containing disulfide bridges.
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18. |
- Banerjee, Debapriya, et al.
(author)
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Spectroscopic and DFT studies on 6-Aminophenanthridine and its derivatives provide insights in their activity towards ribosomal RNA
- 2014
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In: Biochimie. - : Elsevier BV. - 0300-9084 .- 1638-6183. ; 97, s. 194-199
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Journal article (peer-reviewed)abstract
- 6-Aminophenanthridine (6AP), a plant alkaloid possessing antiprion activity, inhibits ribosomal RNA dependent protein folding activity of the ribosome (referred as PFAR). We have compared 6AP and its three derivatives 6AP8Cl, 6AP8CF3 and 6APi for their activity in inhibition of PFAR. Since PFAR inhibition requires 6AP and its derivatives to bind to the ribosomal RNA (rRNA), we have measured the binding affinity of these molecules to domain V of 23S rRNA using fluorescence spectroscopy. Our results show that similar to the antiprion activity, both the inhibition of PFAR and the affinity towards rRNA follow the order 6AP8CF3 > 6AP8Cl > 6AP, while 6APi is totally inactive. To have a molecular insight for the difference in activity despite similarities in structure, we have calculated the nucleus independent chemical shift using first principles density functional theory. The result suggests that the deviation of planarity in 6APi and steric hindrance from its bulky side chain are the probable reasons which prevent it from interacting with rRNA. Finally, we suggest a probable mode of action of 6AP, 6AP8CF3 and 6AP8Cl towards rRNA.
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20. |
- Bartish, Galyna, et al.
(author)
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Importance of individual amino acids in the Switch I region in eEF2 studied by functional complementation in S. cerevisiae
- 2008
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In: Biochimie. - : Elsevier BV. - 0300-9084 .- 1638-6183. ; 90:5, s. 736-748
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Journal article (peer-reviewed)abstract
- Elongation factor 2 (eEF2) is a member of the G-protein super family. G-proteins undergo conformational changes associated with binding of the guanosine nucleotide and hydrolysis of the bound GTP. These structural rearrangements affects the Switch I region (also known as the Effector loop). We have studied the role of individual amino acids in the Switch I region (amino acids 25-73) of S. cerevisiae eEF2 using functional complementation in yeast. 21 point mutations in the Switch I region were created by site-directed mutagenesis. Mutants K49R, E52Q, A53G, F55Y, K60R, Q63A, T68S, 169M and A73G were functional while mutants R54H, F55N, D57A, D57E, D57S, R59K, R59M, Q63E, R65A, R65N, T68A and T68M were inactive. Expression of mutants K49R, A53G, Q63A, 169M and A73G was associated with markedly decreased growth rates and yeast cells expressing mutants A53G and 169M became temperature sensitive. The functional capacity of eEF2 in which the major part Switch I (amino acids T56 to 169) was converted into the homologous sequence found in EF-G from E. coli was also studied. This protein chimera could functionally replace yeast eEF2 in vivo. Yeast cells expressing this mutant grew extremely slowly, showed increased cell death and became temperature sensitive. The ability of the mutant to replace authentic eEF2 in vivo indicates that the structural rearrangement of Switch I necessary for eEF2 function is similar in eukaryotes and bacteria. The effect of two point mutations in the P-loop was also studied. Mutant A25G but not A25V could functionally replace yeast eEF2 even if cells expressing the mutant grew slowly. The A25G mutation converted the consensus sequences AXXXXGK[T/S] in eEF2 to the corresponding motif GXXXXGK[T/S] found in all other G-proteins, suggesting that the alanine found in the P-loop of peptidyltranslocases are not essential for function.
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22. |
- Bjorkhem, I
(author)
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Five decades with oxysterols
- 2013
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In: Biochimie. - : Elsevier BV. - 1638-6183 .- 0300-9084. ; 95:3, s. 448-454
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Journal article (peer-reviewed)
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24. |
- Burchacka, Ewa, et al.
(author)
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Substrate profiling of Finegoldia magna SufA protease, inhibitor screening and application to prevent human fibrinogen degradation and bacteria growth in vitro.
- 2014
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In: Biochimie. - : Elsevier BV. - 1638-6183 .- 0300-9084. ; 103C:May 22, s. 137-143
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Journal article (peer-reviewed)abstract
- SufA, which belongs to the subtilisin-like serine protease family, contains a non-canonical Asp-His-Ser catalytic triad. Under in vitro conditions, SufA is capable of human fibrinogen hydrolysis leading to inhibition of fibrin network formation, thus suggesting its important role in the development and progression of Finegoldia magna infections. In addition, it has been demonstrated that SufA can hydrolyze antibacterial peptides such as LL-37 and the chemokine MIG/CXCL 9, hence evading host defence mechanisms. Although the SufA protease from F. magna was discovered several years ago, its optimal substrate preference has not yet been identified. Considering the role of SufA, we have focused on the profiling of its substrate sequence preference spanning S1-S3 binding pockets using the FRET (fluorescence resonance energy transfer) approach. Next, based on the structure of the P1 residue of the developed substrate, we narrowed the inhibitor screening to the phosphonic analogues of amino acids containing an arginine-like side chain. Among all the compounds tested, only Cbz-6-AmNphth(P)(OPh)2 showed any inhibitory activity against SufA displaying k2/Ki value of 10 800 M(-1) s(-1). In addition, it prevented SufA-mediated human fibrinogen hydrolysis in vitro and exhibited potent antibacterial activity against F. magna, Staphylococcus aureus and Escherichia coli. Herein, we report on the substrate specificity, synthesis and kinetic evaluation of phosphonic inhibitors of SufA protease from F. magna which could help to establish its function in pathogenesis development and may lead to the elaboration of new antibacterial drugs.
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