New convenient reagent for peptide syntheses

Belleau, B.; Malek, G.

doi:10.1021/ja01008a045

Cited by 442 publications

(170 citation statements)

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“…Under these conditions, P-type ATPases quite often are isolated as high molecular mass aggregates by density centrifugation (2,11,14,17,25). In our hands, treatment of 3-14-solubilized ATPase with DOC did not alter the profile of ATPase activity and protein after glycerol gradient centrifugation in the presence of DOC with or without [3][4][5][6][7][8][9][10][11][12][13][14] (data not shown).…”

Section: Characterization Of 3-14-solubilized Atpasementioning

confidence: 68%

“…Microsomes were suspended in 5 mL of homogenizing buffer and layered over a gradient consisting of two 10-mL steps of 34 and 42% (w/w) sucrose in 5 mm Hepes (pH 7.5). After centrifuging for 125 min at 40,000 rpm in a Ti 70 rotor at 40C, the membranes collecting at the interface between the 34 and 42% (w/w) sucrose steps were removed, diluted 3-to 4-fold with 5 When the 3-14-solubilized protein was fractionated by glycerol gradient density centrifugation, the solution was diluted with an equal volume of 5 mm Hepes (pH 7.8). The diluted protein solution was layered over an 11-mL linear gradient of 15 to 40% (w/v) glycerol in 5 mm Hepes (pH 7.8) containing 4 mg/mL of 3-14 either in the presence or absence of 0.2% (w/v) DOC.…”

Section: Isolation Of Plasma Membranesmentioning

confidence: 99%

“…The assay media contained 20 mi Hepes (pH 7.5), 100 mm NaCl, 25 mM KCl, 5 Where indicated, the EEDQ treatment was terminated by diluting the vesicles 5-to 10-fold with ice-cold 20 mm Hepes resuspension buffer. The vesicles were collected after centrifuging at 55,000 rpm in a TLA-100 rotor for 60 min at 40C.…”

Section: Isolation Of Plasma Membranesmentioning

confidence: 99%

“…3), implying that 1 mol of EEDQ inhibited 1 mol of solubilized ATPase. Therefore, solubilization of the corn root P-type H+-ATPase with [3][4][5][6][7][8][9][10][11][12][13][14] corresponded to an alteration in the characteristics of EEDQ inhibition that was consistent with a change in subunit composition.…”

Section: Characteristics Of Eedq Inhibitionmentioning

confidence: 99%

“…EEDQ is able to catalyze the formation of peptide bonds within and between polypeptides by activating carboxylic acids and allowing them to react with nearby amines (4,5). As such, EEDQ has been found to be a potent inhibitor of other P-type ATPases (1).…”

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confidence: 99%

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Effects of Solubilization on the Inhibition of the P-Type ATPase from Maize Roots by N-(Ethoxycarbonyl)-2-Ethoxy-1,2-Dihydroquinoline

Bräuer

Gurriel²,

Tu³

1992

Plant Physiol.

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The biochemical events utilized by transport proteins to convert the chemical energy from the hydrolysis of ATP into an electrochemical gradient are poorly understood. The inhibition of the plasma membrane ATPase from corn (Zea mays L.) roots by N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline (EEDQ) was compared to that of ATPase solubilized with N-tetradecyl-N,N-dimethyl-3-ammonio-1-propane-sulfonate (3-14) to provide insight into the minimal functional unit. The chromatographic behavior of the 3-14-solubilized ATPase activity during size exclusion chromatography and glycerol gradient centrifugation indicated that the solubilized enzyme was in a monomeric form. Both plasma membrane-bound and solubilized ATPase were inhibited by EEDQ in a time-and concentration-dependent manner consistent with a firstorder reaction. When the log of the reciprocal of the half-time for inhibition was plotted as a function of the log of the EEDQ concentration, straight lines were obtained with slopes of approximately 0.5 and 1.0 for membrane-bound and 3-14-solubilized ATPase, respectively, indicating a change in the number of polypeptides per functional ATPase complex induced by solubilization with 3-14.Enzymes that link the transport of ions to either the hydrolysis or synthesis of ATP have been classified into three broad categories of transport ATPase, designated as P-, V-, and F-type (28). These ATPases can be distinguished by their biochemical, physical, and physiological properties. The Ftype ATPase couples the flux of ions, usually protons, down their electrochemical gradient to the synthesis of ATP. The other two classes of ATPases couple the transport of ions against their electrochemical gradient to the release of free energy from ATP hydrolysis.The mechanism by which the energy from ATP is converted into the movement of ions by the V-and P-type ATPases is poorly understood. A generalized reaction scheme has been proposed for the P-type ATPases in which transport reactions are tightly coupled to the reactions, leading to phosphorylation and dephosphorylation of the enzyme (24,28). Soon after the discovery of the V-type ATPase, it was apparent that a reaction mechLanism in which ATP hydrolysis and ion transport were tightly linked could not account for the differential inhibition and activation of the activities of

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Section: Characterization Of 3-14-solubilized Atpasementioning

confidence: 68%

Section: Isolation Of Plasma Membranesmentioning

confidence: 99%

Section: Isolation Of Plasma Membranesmentioning

confidence: 99%

Section: Characteristics Of Eedq Inhibitionmentioning

confidence: 99%

mentioning

confidence: 99%

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Effects of Solubilization on the Inhibition of the P-Type ATPase from Maize Roots by N-(Ethoxycarbonyl)-2-Ethoxy-1,2-Dihydroquinoline

Bräuer

Gurriel²,

Tu³

1992

Plant Physiol.

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Inhibition of cysteine proteases by peptides containing aziridine-2,3-dicarboxylic acid building blocks

Schirmeister

1999

Biopolymers

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A pH- and ionic strength-responsive polypeptide hydrogel: Synthesis, characterization, and preliminary protein release studies

Markland

Zhang

Amidon

et al. 1999

J. Biomed. Mater. Res.

122

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A novel polypeptide hydrogel has been synthesized by crosslinking poly(L-glutamic acid) (PLG) with poly(ethylene glycol) (PEG). The PLG-PEG hydrogel was shown to be highly hydrophilic, and the extent of swelling varied with pH, increasing at higher ionization of the PLG. Aside from electrostatic effects, such as ion-ion repulsion and internal ion osmotic pressure, circular dichroism studies showed that swelling response to pH also is affected by secondary structural attributes associated with the polypeptide backbone. Modification of the polypeptide by changing its hydrophobicity and degree of ionization was an effective method for altering the overall extent of pH-responsive swelling. Rapid de-swelling (contraction) was observed when the PLG-PEG hydrogel was transferred from high to low pH buffer solution, and this swelling/de-swelling behavior was reversible over repeated cycles. Drug release from swollen hydrogels was examined using the model protein lysozyme. Rapid de-swelling of the hydrogel was found to be an effective means of facilitating lysozyme release. The crosslinking of synthetic polypeptides with PEG appears to be a highly versatile approach to the preparation of pH-responsive biodegradable hydrogels.

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New convenient reagent for peptide syntheses

Cited by 442 publications

References 1 publication

Effects of Solubilization on the Inhibition of the P-Type ATPase from Maize Roots by N-(Ethoxycarbonyl)-2-Ethoxy-1,2-Dihydroquinoline

Effects of Solubilization on the Inhibition of the P-Type ATPase from Maize Roots by N-(Ethoxycarbonyl)-2-Ethoxy-1,2-Dihydroquinoline

Inhibition of cysteine proteases by peptides containing aziridine-2,3-dicarboxylic acid building blocks

A pH- and ionic strength-responsive polypeptide hydrogel: Synthesis, characterization, and preliminary protein release studies

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