Sensitivity of phospholipase C (Bacillus cereus) activity to phosphatidylcholine structural modifications

El-Sayed, M.Y.; DeBose, Cheryl D.; Coury, Larry A.; Roberts, Mary F.

doi:10.1016/0005-2760(85)90056-6

Cited by 80 publications

(49 citation statements)

References 19 publications

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“…290 for PLA # and approx. 11 for PLC, extends well beyond that observed for aliphatic PCs, and (2) the optimal length for the fatty acyl chain at sn-2 in BHPCs is 16 atoms (BH-C12PC), whereas dioctanoyl PC (or diheptanoyl PC for PLC) [9,10] is the best substrate in the aliphatic series.…”

Section: Structure Of Bhpcs Influences the Hydrolytic Activity Of Phomentioning

confidence: 54%

“…3-(4-Acetoxyphenyl)propionic acid (9) Compound 8 oBolton-Hunter [BH, 3-(4-hydroxyphenyl)-propionic acid] 400 mg, 2.4 mmol, UV positive spot of R F 0.22 by TLC developed in solvent 3q (see Scheme 1) and acetic anhydride (250 mg, 3.37 mmol) were dissolved in anhydrous chloroform (10 ml) and the mixture was cooled to 0 mC. The reaction was catalysed by the addition of a drop of concentrated (18 M) H # SO % .…”

Section: Synthesis and Characterization Of Compoundsmentioning

confidence: 99%

“…aggregated substrates, i.e. above the critical micelle concentration [8,9]. Several studies have defined the structural requirements for phospholipids to become substrates of phospholipases [9][10][11].…”

Section: Introductionmentioning

confidence: 99%

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Mapping the catalytic pocket of phospholipases A2 and C using a novel set of phosphatidylcholines

Caramelo

Florin‐Christensen

Florín-Christensen

et al. 2000

Biochemical Journal

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A set of radioiodinatable phosphatidylcholines (PCs) derivatized with the Bolton-Hunter reagent (BHPCs) was synthesized to probe the substrate recognition and activity of phospholipases. A common feature of this series is the presence of a bulky 4-hydroxyphenyl group at the end of the fatty acyl chain attached to position sn-2. The distance between the end group and the glycerol backbone was varied by changing the length of the intervening fatty acyl chain (3-25 atoms). Except for the shortest, this chain includes at least one amide linkage. The usefulness of this series of substrates as a molecular ruler was tested by measuring the hydrolytic activities of Naja naja naja phospholipase A2 (PLA2) and Bacillus cereus phospholipase C (PLC) in Triton X-100 micelles. The activity of PLA2 proved to be highly dependent on the length of the fatty acyl chain linker, the shorter compounds (3-10 atoms) being very poor substrates. In contrast, the PLC activity profile exhibited much less discrimination. In both cases, PCs with 16-21 atom chains at position sn-2 yielded optimal activity. We interpret these findings in terms of fatty acyl chain length-related steric hindrance caused by the terminal aromatic group, affecting the activity of PLA2 and, to a smaller extent, that of PLC. This notion agrees with the more extended recognition of aliphatic chains inside the narrow channel leading to the catalytic site in the former case. Molecular models of these substrates bound to PLA2 were built on the basis of the crystallographic structure of Naja naja atra PLA2 complexed with a phospholipid analogue. Docking of these substrates necessarily requires the intrusion of the bulky 4-hydroxyphenyl group inside the binding pocket and also the failure of the amide group to form hydrogen bonds inside the hydrophobic substrate channel.

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Section: Structure Of Bhpcs Influences the Hydrolytic Activity Of Phomentioning

confidence: 54%

Section: Synthesis and Characterization Of Compoundsmentioning

confidence: 99%

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Mapping the catalytic pocket of phospholipases A2 and C using a novel set of phosphatidylcholines

Caramelo

Florin‐Christensen

Florín-Christensen

et al. 2000

Biochemical Journal

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“…Among the three major regions in the membrane, the interfacial region is characterized by unique motional and dielectric characteristics (Ashcroft et al, 1981) different from the bulk aqueous phase (experienced by charged aqueous probes such as ANS and TNS) and the more isotropic hydrocarbon-like deeper regions of the membrane and plays an important role in functional aspects such as substrate recognition and activity of lipolytic enzymes (El-Sayed et al, 1985). This specific region of the membrane exhibits slow rates of solvent relaxation and is also known to participate in intermolecular charge interactions (Yeagle, 1987) and hydrogen bonding through the polar headgroup (Boggs, 1987;Gennis, 1989;Shin et al, 1991).…”

Section: The Wavelength-selective Fluorescence Approachmentioning

confidence: 99%

Exploring membrane organization and dynamics by the wavelength-selective fluorescence approach

Chattopadhyay

2003

Chemistry and Physics of Lipids

144

152

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“…The NBD group in NBD-PE is located at the membrane interface. This region of the membrane has unique motional and dielectric characteristics distinct from both the bulk aqueous phase and the hydrocarbon-like interior of the membrane (Ashcroft et al, 1981;Stubbs et al, 1985;Perochon et al, 1992) and plays an important role in functional aspects such as substrate recognition and activity of lipolytic enzymes (El-Sayed et al, 1985). Application of the red edge effect to probes whose location in membranes are well defined could provide useful information regarding the structural organization and dynamics in specific regions of the membrane.…”

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confidence: 99%

Fluorophore environments in membrane-bound probes: A red edge excitation shift study

Chattopadhyay

Mukherjee²

1993

Biochemistry

140

158

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A shift in the wavelength of maximum fluorescence emission toward higher wavelengths, caused by a shift in the excitation wavelength toward the red edge of the absorption band, is termed the Red Edge Excitation Shift (REES). This effect is mostly observed with polar fluorophores in motionally restricted media such as very viscous solutions or condensed phases. In this paper, we report the red edge excitation shift of a membrane-bound phospholipid molecule whose headgroup is covalently labeled with a 7-nitrobenz-2-oxa-1,3-diazol-4-y1 (NBD) moiety. When incorporated into model membranes of dioleoyl-sn-glycero-3-phosphocholine (DOPC), the NBD-labeled phospholipid (NBD-PE), exhibits a red edge excitation shift of 10 nm. In addition, fluorescence polarization of NBD-PE in membranes shows both excitation and emission wavelength dependence. The nonpolar membrane probe 1,6-diphenyl-1,3,5-hexatriene (DPH) does not show red edge excitation shift in model membranes. The lifetime of NBD-PE in DOPC vesicles was found to be dependent on both excitation and emission wavelengths. These wavelength-dependent lifetimes are correlated to the reorientation of solvent dipoles around the excited-state dipole of the NBD moiety in the membrane. The magnitude of the red shift in the emission maximum for NBD-PE was found to be independent of temperature, between 12 and 54 O C , and of the physical state (gel or fluid) of the membrane. Taken together, these observations are indicative of the motional restriction experienced by this fluorophore in the membrane. Red edge excitation shift promises to be a powerful tool in probing membrane organization and dynamics.Organized molecular assemblies such as membranes can be considered as large cooperative units with characteristics very different from the individual structural units which constitute them. Fluorescence spectroscopy has been one of the principal techniques to study organization and dynamics of biological and model membranes because of its suitable time scale, noninvasive nature, and intrinsic sensitivity (Radda, 1975;Lakowicz, 1980Lakowicz, , 1981. For a fluorophore in a bulk nonviscous solvent, the dipolar relaxation of the solvent molecules around the fluorophore in the excited state is much faster than the fluorescence lifetime. The fluorescence decay rates and the wavelength of maximum emission, in such a case, are therefore usually independent of the excitation wavelength. However, these parameters do show excitation wavelength dependence if the dipolar relaxation of the solvent molecules is slow in the excited state, such that the relaxation time is comparable to or longer than the fluorescence lifetime (Chen, 1967;Fletcher, 1968;Galley & Purkey, 1970;Rubinov & Tomin, 1970;Castelli & Forster, 1973;Itoh & Azumi, 1975;Demchenko, 1982; Lakowicz & Keating-Nakamoto, 1984). Such a shift in the wavelength of maximum emission toward higher wavelengths, caused by a shift in the excitation wavelength toward the red edge of the absorption band, is termed the red edge excitatio...

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Sensitivity of phospholipase C (Bacillus cereus) activity to phosphatidylcholine structural modifications

Cited by 80 publications

References 19 publications

Mapping the catalytic pocket of phospholipases A2 and C using a novel set of phosphatidylcholines

Mapping the catalytic pocket of phospholipases A2 and C using a novel set of phosphatidylcholines

Exploring membrane organization and dynamics by the wavelength-selective fluorescence approach

Fluorophore environments in membrane-bound probes: A red edge excitation shift study

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