INTERFACIAL ENZYMOLOGY OF GLYCEROLIPID HYDROLASES: Lessons from Secreted Phospholipases A<sub>2</sub>

Gelb, Michael H.; Jain, Mahendra Kumar; Hanel, Arthur M.; Berg, Otto G.

doi:10.1146/annurev.bi.64.070195.003253

Cited by 233 publications

(234 citation statements)

References 120 publications

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“…Catalytically active hGXII was produced in Escherichia coli and shown to be Ca 2؉ -dependent despite the fact that it is predicted to have an unusual Ca 2؉ -binding loop. Similar to the previously characterized mouse group IIE sPLA 2 s, the specific activity of hGXII is low in comparison to that of other mammalian sPLA 2 , suggesting that hGXII could have novel functions that are independent of its phospholipase A 2 activity.…”

supporting

confidence: 72%

Cloning and Recombinant Expression of a Structurally Novel Human Secreted Phospholipase A2

Gelb¹,

Valentin²,

Ghomashchi³

et al. 2000

Journal of Biological Chemistry

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161

152

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Mammals contain a diverse set of secreted phospholipases A 2 (sPLA 2 s) that liberate arachidonic acid from phospholipids for the production of eicosanoids and exert a variety of physiological and pathological effects. We report the cloning, recombinant expression, and kinetic properties of a novel human sPLA 2 that defines a new structural class of sPLA 2 s called group XII. The human group XII (hGXII) cDNA contains a putative signal peptide of 22 residues followed by a mature protein of 167 amino acids that displays homology to all known sPLA 2 s only over a short stretch of amino acids in the active site region. Northern blot and reverse transcription-polymerase chain reaction analyses show that the tissue distribution of hGXII is distinct from the other human sPLA 2 s with strong expression in heart, skeletal muscle, kidney, and pancreas and weaker expression in brain, liver, small intestine, lung, placenta, ovaries, testis, and prostate. Catalytically active hGXII was produced in Escherichia coli and shown to be Ca 2؉ -dependent despite the fact that it is predicted to have an unusual Ca 2؉ -binding loop. Similar to the previously characterized mouse group IIE sPLA 2 s, the specific activity of hGXII is low in comparison to that of other mammalian sPLA 2 , suggesting that hGXII could have novel functions that are independent of its phospholipase A 2 activity.

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supporting

confidence: 72%

Cloning and Recombinant Expression of a Structurally Novel Human Secreted Phospholipase A2

Gelb¹,

Valentin²,

Ghomashchi³

et al. 2000

Journal of Biological Chemistry

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161

152

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“…However, K132 was replaced with glutamic acid (and not with glutamine residue, which is present in AtnI # ), to lower the pI of the sextuple AtxA mutant. Charge reversal of ammodytoxin A Figure 1 Sequence alignment of basic, neurotoxic ammodytoxin A with neutral, non-toxic AtnI 2 The common numbering of PLA 2 residues is used [27]. The substitutions in the sextuple and single mutants of AtxA are shown by arrows.…”

Section: Resultsmentioning

confidence: 99%

Charge reversal of ammodytoxin A, a phospholipase A2-toxin, does not abolish its neurotoxicity

Prijatelj

Čopič

Križaj

et al. 2000

Biochemical Journal

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The positive charge concentrated at the C-terminal region of ammodytoxin (Atx) A, which is involved in presynaptic toxicity, has been reversed. A six-site mutant of AtxA (K108N/K111N/K127T/K128E/E129T/K132E, where K108N = Lys108 → Asn etc.) was prepared, in which five out of seven C-terminal basic amino acid residues were substituted with neutral or acidic ones. The mutant was approximately 30-fold less lethal, but still neurotoxic. Consistent with this, its binding affinity for the neuronal receptors decreased by only a factor of five. Additionally, a single-site mutant of AtxA was prepared, with substitution at only one position (K127T) out of six mutated in the six-site mutant. Its toxicity indicated that most, if not all, of the six mutated residues partially contribute to the decreased lethality of the multiple-site mutant.

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“…Both the viral and mammalian PLA 2 s are able to readily hydrolyze phosphatidylcholine vesicles (Table II), and thus it was expected that the viral PLA 2 s could act on the outer monolayer of mammalian cells, which is rich in zwitterionic phospholipids. In contrast, human group IIA sPLA 2 binds weakly to phosphatidylcholine-rich membranes and fails to release arachidonic acid when added exogenously to mammalian cells at concentrations up to 1 g/ml (16). The intact PPV virus particle undergoes a structural transition after heat shock that greatly enhances the PLA 2 enzymatic activity and externalizes the VP1up PLA 2 domain (Fig.…”

Section: Discussionmentioning

confidence: 99%

Interfacial Enzymology of Parvovirus Phospholipases A2

Canaan

Zádori

Ghomashchi

et al. 2004

Journal of Biological Chemistry

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105

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The capsid of parvoviruses proteins were recently shown to contain secreted phospholipase A 2 (sPLA 2 )-like activity that is required during host cell entry. Parvoviral PLA 2 domains have little sequence identity with sPLA 2 s and lack disulfide bonds. In the present study, after bacterial expression and purification, the biochemical characterizations of these first PLA 2 s identified in viruses have been investigated, and a comparison has been made with other known PLA 2 s. The specific activities of three viral PLA 2 s differed by 3 orders of magnitude, with porcine parvovirus PLA 2 displaying a specific activity similar to that of the most active sPLA 2 s (e.g. human group IIA) and the human AAV2 and B19 parvoviral enzymes displaying approximately 10 3 lower specific activities (similar to human sPLA 2 groups IIE and XIIA). These differences were not caused by weaker Ca 2؉ or interfacial binding. The specific activities of the viral PLA 2 s on zwitterionic or anionic phospholipid vesicles were comparable. The viral PLA 2 s did not display a preference for unsaturated versus saturated sn-2 fatty acyl chains and hydrolyzed all major classes of glycerophospholipids except phosphatidylinositol. Incubation of mammalian cells with porcine parvovirus PLA 2 led to the release of arachidonic acid into the culture medium. Interestingly, among nine previously known sPLA 2 inhibitors, only a subset showed inhibition of the viral PLA 2 s and with weak potency, indicating that the active sites of these new enzymes are structurally distinct from those of sPLA 2 s. Based on these distinct enzymatic and structural properties, we propose to classify the parvovirus PLA 2 s within the PLA 2 superfamily as group XIII enzymes.Parvoviruses are a family of small DNA viruses comprising two subfamilies, the Parvovirinae that infect vertebrates, including man, and the Densovirinae that infect invertebrates such as insect and shrimp species. These viruses depend, because of their limited genome content, on enzymes and precursors present during the S phase of the cell for their replicative functions and thus show a proclivity for actively dividing cells. Consequently, many parvoviruses, such as the porcine parvovirus (1) (PPV) 1 and the human B19 (2) parvovirus, are fetotropic, resulting in abortions, or have a predisposition to infect other mitosis-active tissues such as transformed cells ("oncolytic property") (3).These nonenveloped, icosahedral viruses have a diameter of approximately 25 nm and contain a linear, single-stranded DNA genome of approximately 4 -6 kb. The structure of several parvovirus capsids has been solved to nearly atomic resolution, whereas the DNA is only partly ordered (4, 5). The capsid is composed of 60 structurally equivalent protein subunits, each containing a ␤-barrel jelly roll in which some loops between the ␤-strands intertwine with loops from neighboring subunits and cover the viral surface for specific functions such as recognition of cellular receptors. Approximately 10 out of the 60 capsid proteins ha...

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INTERFACIAL ENZYMOLOGY OF GLYCEROLIPID HYDROLASES: Lessons from Secreted Phospholipases A₂

Cited by 233 publications

References 120 publications

Cloning and Recombinant Expression of a Structurally Novel Human Secreted Phospholipase A2

Cloning and Recombinant Expression of a Structurally Novel Human Secreted Phospholipase A2

Charge reversal of ammodytoxin A, a phospholipase A2-toxin, does not abolish its neurotoxicity

Interfacial Enzymology of Parvovirus Phospholipases A2

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INTERFACIAL ENZYMOLOGY OF GLYCEROLIPID HYDROLASES: Lessons from Secreted Phospholipases A2

Cited by 233 publications

References 120 publications

Cloning and Recombinant Expression of a Structurally Novel Human Secreted Phospholipase A2

Cloning and Recombinant Expression of a Structurally Novel Human Secreted Phospholipase A2

Charge reversal of ammodytoxin A, a phospholipase A2-toxin, does not abolish its neurotoxicity

Interfacial Enzymology of Parvovirus Phospholipases A2

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INTERFACIAL ENZYMOLOGY OF GLYCEROLIPID HYDROLASES: Lessons from Secreted Phospholipases A₂