Polymerizable phosphatidylcholines: Importance of phospholipid motions for optimum phospholipase A2 and C activity

Soltys, Christine E.; Bian, Jinru; Roberts, Mary F.

doi:10.1021/bi00088a005

Cited by 24 publications

(27 citation statements)

References 28 publications

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“…Nevertheless, the putative entrance channel for the substrate in PLC is broader and shallower than that in PLA # , bringing about a less pronounced activity pattern for the former enzyme. Constraints on the vertical movement of the substrate are less stringent in PLC [12,37], as documented here by the behaviour of BH-C20PC, for which the increased chain length caused only a minor detrimental effect on enzymic activity.…”

Section: Structure Of Bhpcs Influences the Hydrolytic Activity Of Phomentioning

confidence: 78%

“…In any of these models, an impairment of optimal catalytic efficiency will result. In this context, it is known that a constraint on the vertical movement of the substrate negatively affects the activity of phospholipases [12,37].…”

Section: Structure Of Bhpcs Influences the Hydrolytic Activity Of Phomentioning

confidence: 99%

“…Besides, no impediment should preclude the upward motion of the substrate from the lipid phase into the catalytic site, e.g. by anchoring the tail of the fatty acid to a polymeric support [12,13]. More generally, any structural feature of the substrate that restricts the vertical movement of the molecule markedly affects enzymic activity.…”

Section: Introductionmentioning

confidence: 99%

“…More generally, any structural feature of the substrate that restricts the vertical movement of the molecule markedly affects enzymic activity. In contrast, PLC is less stringent than PLA # in its substrate requirements, the chain length dependence is not so important, and a constraint on the vertical motion of the substrate does not seem to affect enzymic activity negatively [12,13]. In the light of the structures of several phospholipases, crystallized in their isolated forms or in complexes with substrate analogues [14][15][16][17], the existence of hydrophobic channels leading to the catalytic sites was proposed.…”

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: 78%

Section: Structure Of Bhpcs Influences the Hydrolytic Activity Of Phomentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

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 commonly accepted model emerging from those studies is that 1) the enzyme associates peripherally with the macrosubstrate surface, 2) the phospholipid substrate moves upward and engages with the active site cavity of the enzyme, and 3) hydrolysis of the sn2 ester bond takes place, and the products are released from the enzyme (8,18,19,(45)(46)(47)(48)(49)(50). Based on this model, the key factors determining the substrate specificity can be considered to be 1) accommodation of phospholipid acyl chain(s) in the active site cavity of the enzyme and 2) propensity of the phospholipid substrate to efflux from the bilayer.…”

Section: Discussionmentioning

confidence: 99%

Substrate Efflux Propensity Plays a Key Role in the Specificity of Secretory A-type Phospholipases

Haimi¹,

Hermansson²,

Batchu³

et al. 2010

Journal of Biological Chemistry

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To better understand the principles underlying the substrate specificity of A-type phospholipases (PLAs), a high throughput mass spectrometric assay was employed to study the effect of acyl chain length and unsaturation of phospholipids on their rate of hydrolysis by three different secretory PLAs in micelles and vesicle bilayers. With micelles, each enzyme responded differently to substrate acyl chain unsaturation and double bond position, probably reflecting differences in the accommodative properties of their substrate binding sites. Experiments with saturated acyl positional isomers indicated that the length of the sn2 chain was more critical than that of the sn1 chain, suggesting tighter association of the former with the enzyme. Only the first 9 -10 carbons of the sn2 acyl chain seem to interact intimately with the active site. Strikingly, no discrimination between positional isomers was observed with vesicles, and the rate of hydrolysis decreased far more with increasing chain length than with micelles, suggesting that translocation of the phospholipid substrate to the active site is rate-limiting with bilayers. Supporting this conclusion, acyl chain structure affected hydrolysis and spontaneous intervesicle transfer, which correlates with lipid efflux propensity, analogously. We conclude that substrate efflux propensity plays a more important role in the specificity of secretory PLA 2 s than commonly thought and could also be a key attribute in phospholipid homeostasis in which (unknown) PLA 2 s are key players.Type A phospholipases (PLAs) 3 are ubiquitous enzymes involved in many biological phenomena, such as signal transduction (1) and phospholipid homeostasis (2), including acyl chain remodeling (3) and degradation (4). However, the identities of the specific proteins involved in these phenomena have often not been established. PLAs are also clinically relevant because they have been implicated in many common diseases or disorders, including atherosclerosis, allergy, Alzheimer disease, and cancer (5).Several PLAs display significant specificity toward the phospholipid acyl chain(s) in vitro (6, 7). However, despite numerous studies, the factors underlying such specificity have remained largely elusive. In principle, two factors contribute to selective hydrolysis of phospholipids by PLA: 1) accommodation of the acyl chains in the protein lipid binding site and 2) the ease of efflux of the phospholipid substrate from the bilayer (8, 9). The understanding of acyl chain accommodation has been greatly hampered by the lack of crystal structures of PLAs complexed with a physiological substrate. Crystal structures have been obtained for some PLAs with a bound noncleavable shortchain substrate analogue (10 -13), but because these analogues have truncated acyl chains, they only provide very limited information on the factors underlying acyl chain specificity. Furthermore, the information could also be biased, since studies with other lipid-binding proteins have shown that the mode of accommodation of an alkyl...

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Synthesis of terpolymers having a phospholipid polar group and poly(oxyethylene) in the side chain and their protein adsorption–resistance properties

Oishi

Tanaka

Yamasaki

et al. 2002

J of Applied Polymer Sci

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Terpolymers having phospholipid polar groups were synthesized from 2-methacryloyloxyethyl phosphorylcholine (MPC), methacryloyl or acryloyl poly(oxyethylene) macromonomers (POEM) [(CH 2 CH 2 O) n (where n ϭ 2-23); PEOM(2), PEOM(23), ME(9), Ph(6)], and n-butyl methacrylate (BMA). The characteristics of these terpolymer membranes were investigated by water content (H) and X-ray photoelectron spectroscopy. The content of water in the terpolymer increased with increasing content of MPC and length of oxyethylene units. The membranes of terpolymers were found to adsorb bovine serum albumin much less than those of poly(methyl methacrylate) and poly(BMA). Even though the contents of MPC in the terpolymer were 5 to 25 mol %, the terpolymer depressed BSA adsorption more than poly(MPC-co-BMA) consisting of 29 mol % of MPC. The use of terpolymer with POEM can decrease the amount of MPC in the polymer.

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Polymerizable phosphatidylcholines: Importance of phospholipid motions for optimum phospholipase A2 and C activity

Cited by 24 publications

References 28 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

Substrate Efflux Propensity Plays a Key Role in the Specificity of Secretory A-type Phospholipases

Synthesis of terpolymers having a phospholipid polar group and poly(oxyethylene) in the side chain and their protein adsorption–resistance properties

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