Ion-exchange properties of cibacron blue 3G-A sepharose (blue sepharose) and the interaction of proteins with cibacron blue 3g-a

Lascu, Ioan; Porumb, Horea; Porumb, Tudor; Abrudan, I.; Tărmure, C; Petrescu, Ioan; Presecan, Elena; Proinov, Ioan; Telia, M.

doi:10.1016/s0021-9673(00)96255-1

Cited by 34 publications

(11 citation statements)

References 20 publications

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“…Previous studies have also revealed comparable arrangements with cytochrome C, [ 23 ] which is almost entirely α-helical. [ 27 ] In fact, the density of the amphiphilic molecules at the interface plays a key role in determining the orientation of the liquid crystal with which they are in contact. [ 11,16,17 ] When lipid monolayers at low lipid densities are transferred to the aqueous-LC interface, a tilted orientation is observed.…”

Section: Resultsmentioning

confidence: 99%

Liquid Crystal Enabled Early Stage Detection of Beta Amyloid Formation on Lipid Monolayers

Sadati

Apik

Armas-Pérez

et al. 2015

Adv Funct Materials

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Liquid crystals (LCs) can serve as sensitive reporters of interfacial events, and this property has been used for sensing of synthetic or biological toxins. Here it is demonstrated that LCs can distinguish distinct molecular motifs and exhibit a specific response to beta‐sheet structures. That property is used to detect the formation of highly toxic protofibrils involved in neurodegenerative diseases, where it is crucial to develop methods that probe the early‐stage aggregation of amyloidogenic peptides in the vicinity of biological membranes. In the proposed method, the amyloid fibrils formed at the lipid–decorated LC interface can change the orientation of LCs and form elongated and branched structures that are amplified by the mesogenic medium; however, nonamyloidogenic peptides form ellipsoidal domains of tilted LCs. Moreover, a theoretical and computational analysis is used to reveal the underlying structure of the LC, thereby providing a detailed molecular‐level view of the interactions and mechanisms responsible for such motifs. The corresponding signatures can be detected at nanomolar concentrations of peptide by polarized light microscopy and much earlier than the ones that can be identified by fluorescence‐based techniques. As such, it offers the potential for early diagnoses of neurodegenerative diseases and for facile testing of inhibitors of amyloid formation.

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Section: Resultsmentioning

confidence: 99%

Liquid Crystal Enabled Early Stage Detection of Beta Amyloid Formation on Lipid Monolayers

Sadati

Apik

Armas-Pérez

et al. 2015

Adv Funct Materials

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“…Calmodulin is the eukaryotic protein that binds within a subdomain of the N terminus of B. pertussis ACT and stimulates its adenylate cyclase activity (51,92). To address this affinity issue, we evaluated binding of ACT to Blue-Sepharose, a reactive dye matrix that interacts with proteins through their nucleotide binding sites (54,80). This method has been previously utilized to determine ACT binding affinities based on the differential binding ability of ACT for Blue-Sepharose in the presence and absence of CaM (50).…”

Section: Reverse Transcription-pcr (Rt-pcr)mentioning

confidence: 99%

Adenylate Cyclase Toxin (ACT) fromBordetella hinzii: Characterization and Differences from ACT ofBordetella pertussis

et al. 2005

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Bordetella hinzii is a commensal respiratory microorganism in poultry but is increasingly being recognized as an opportunistic pathogen in immunocompromised humans. Although associated with a variety of disease states, practically nothing is known about the mechanisms employed by this bacterium. In this study, we show by DNA sequencing and reverse transcription-PCR that both commensal and clinical strains of B. hinzii possess and transcriptionally express cyaA, the gene encoding adenylate cyclase toxin (ACT) in other pathogenic Bordetella species. By Western blotting, we also found that B. hinzii produces full-length ACT protein in quantities that are comparable to those made by B. pertussis. In contrast to B. pertussis ACT, however, ACT from B. hinzii is less extractable from whole bacteria, nonhemolytic, has a 50-fold reduction in adenylate cyclase activity, and is unable to elevate cyclic AMP levels in host macrophages (nontoxic). The decrease in enzymatic activity is attributable, at least in part, to a decreased binding affinity of B. hinzii ACT for calmodulin, the eukaryotic activator of B. pertussis ACT. In addition, we demonstrate that the lack of intoxication by B. hinzii ACT may be due to the absence of expression of cyaC, the gene encoding the accessory protein required for the acylation of B. pertussis ACT. These results demonstrate the expression of ACT by B. hinzii and represent the first characterization of a potential virulence factor of this organism.

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“…New batches of affinity material must be tested in order to find the optimal ATP concentration to be used for the elu-tion. Using a weakly substituted gel the nonspecific, ion-exchange binding of other proteins is minimal, especially at pH 8 [23], although a larger gel-to-protein ratio has to be used. A key feature of the affinity experiment was the binding step at a pH above 8.0 (i.e.…”

Section: Resultsmentioning

confidence: 99%

Nucleoside diphosphate kinase from human erythrocytes: Purification, molecular mass and subunit structure

Presecan¹,

Vonica²,

Lascu³

1989

FEBS Letters

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A new procedure for the purification of nucleoside diphosphate kinase from human erythrocytes is described. The enzyme (105 kDa by gel filtration) is made up of two different kinds of subunits (19.0 and 20.5 kDa), both displaying enzymatic activity. The probable subunit structure of the enzyme is hexameric. The discrepancies related to earlier work are discussed.Nucleoside diphosphate kinase; Affinity chromatography; Subunit structure; (Human erythrocyte)

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Ion-exchange properties of cibacron blue 3G-A sepharose (blue sepharose) and the interaction of proteins with cibacron blue 3g-a

Cited by 34 publications

References 20 publications

Liquid Crystal Enabled Early Stage Detection of Beta Amyloid Formation on Lipid Monolayers

Liquid Crystal Enabled Early Stage Detection of Beta Amyloid Formation on Lipid Monolayers

Adenylate Cyclase Toxin (ACT) fromBordetella hinzii: Characterization and Differences from ACT ofBordetella pertussis

Nucleoside diphosphate kinase from human erythrocytes: Purification, molecular mass and subunit structure

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