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PubMed2005

Potato tuber isoapyrases: substrate specificity, affinity labeling, and proteolytic susceptibility.

Apyrase/ATP-diphosphohydrolase hydrolyzes di- and triphosphorylated nucleosides in the presence of a bivalent ion with sequential release of orthophosphate. We performed studies of substrate specificity on homogeneous isoapyrases from two potato tuber clonal varieties: Desiree (low ATPase/ADPase ratio) and Pimpernel (high ATPase/ADPase ratio) by measuring the kinetic parameters K(m) and k(cat) on deoxyribonucleotides and fluorescent analogues of ATP and ADP. Both isoapyrases showed a broad specificity towards dATP, dGTP, dTTP, dCTP, thio-dATP, fluorescent nucleotides (MANT-; TNP-; ethene-derivatives of ATP and ADP). The hydrolytic activity on the triphosphorylated compounds was always higher for the Pimpernel apyrase. Modifications either on the base or the ribose moieties did not increase K(m) values, suggesting that the introduction of large groups (MANT- and TNP-) in the ribose does not produce steric hindrance on substrate binding. However, the presence of these bulky groups caused, in general, a reduction in k(cat), indicating an important effect on the catalytic step. Substantial differences were observed between potato apyrases and enzymes from various animal tissues, concerning affinity labeling with azido-nucleotides and FSBA (5'-p-fluorosulfonylbenzoyl adenosine). PLP-nucleotide derivatives were unable to produce inactivation of potato apyrase. The lack of sensitivity of both potato enzymes towards these nucleotide analogues rules out the proximity or adequate orientation of sulfhydryl, hydroxyl or amino-groups to the modifying groups. Both apyrases were different in the proteolytic susceptibility towards trypsin, chymotrypsin and Glu-C.

Affinity Labels↗
PubMed1995

Use of synthetic peptides and copolymers to study the substrate specificity and inhibition of the protein tyrosine kinase pp60c-src.

The ability of synthetic peptides and polypeptides to act as substrates and/or inhibitors of pp60c-src was examined. The random copolymer, poly(K4Y) had a threefold lower specificity than poly(E4Y). Peptides containing lysine vs. glutamate were also found to have a lower substrate specificity (Vmax:Km ratio). In order to assess the substrate specificity of acidic peptides, an assay protocol using DEAE-membranes was developed. Peptides containing a (YXE)5YXD motif (X = G, A, V, P, or norvaline) were tested as inhibitors and substrates of pp60c-src. The glycine-containing peptide was the best substrate having a specificity 16,000-fold higher than 5Val-angiotensin II, the most commonly used peptide substrate. Most of the peptides, except for the proline containing peptide, had Ki values of 20-100 microM. In a series of (XGE)5XGD peptides, where X = Y or F, tyrosine at position 10 was found to be the preferred site for accepting a phosphate. Analogs in which the glycine was replaced with alanine indicated that loss of flexibility around position 10 was detrimental to substrate specificity. Results suggest that conformational requirements of the peptides tested was important and substrate specificity was a more sensitive parameter than binding as measured by Ki values.

Amino Acid Sequence↗
PubMed1996

Fine substrate specificities of four exo-type cellulases produced by Aspergillus niger, Trichoderma reesei, and Irpex lacteus on (1-->3), (1-->4)-beta-D-glucans and xyloglucan.

To investigate the fine substrate specificities of four highly purified exo-type cellulases (Exo-A from Aspergillus niger, CBHI and CBHII from Trichoderma reesei, and Ex-1 from Irpex lacteus), water-soluble substrates such as barley glucan, xyloglucan from tamarind (Tamarindus indica L.), and their oligosaccharides were employed. Four exo-type cellulases immediately hydrolyzed 3-O-beta-D-cellotriosylglucose to produce cellobiose and laminaribiose. In contrast, CBHII showed no hydrolytic activity towards 3(2)-O-beta-D-cello-biosylcellobiose, which was hydrolyzed to cellobiose by the other exo-type cellulases. These cellulases hydrolyzed the internal linkages of barley glucan and lichenan in an endo-type fashion to produce cellobiose and mix-linked oligosaccharides as main products. The DP-lowering activities of the four exo-type cellulases on barley glucan were in the order of Ex-1, CBHII, Exo-A, and CBHI. Based on gel permeation chromatography analysis of the hydrolysates, Ex-1 seemed to attack the internal cellobiosyl unit adjacent to beta-1,3-glucosidic linkages in barley glucan molecule more frequently than did the other cellulases. Xyloglucan was hydrolyzed only by CBHI and CBHII, and produced hepta-, octa-, and nona-saccharides. In addition, a xyloglucan tetradecasaccharide (XG14) was split only to heptasaccharide (XG7) by CBHI and CBHII.

Aspergillus niger↗
PubMed1999

Yeast methionine aminopeptidase I. Alteration of substrate specificity by site-directed mutagenesis.

In eukaryotes, two isozymes (I and II) of methionine aminopeptidase (MetAP) catalyze the removal of the initiator methionine if the penultimate residue has a small radius of gyration (glycine, alanine, serine, threonine, proline, valine, and cysteine). Using site-directed mutagenesis, recombinant yeast MetAP I derivatives that are able to cleave N-terminal methionine from substrates that have larger penultimate residues have been expressed. A Met to Ala change at 329 (Met206 in Escherichia coli enzyme) produces an average catalytic efficiency 1.5-fold higher than the native enzyme on normal substrates and cleaves substrates containing penultimate asparagine, glutamine, isoleucine, leucine, methionine, and phenylalanine. Interestingly, the native enzyme also has significant activity with the asparagine peptide not previously identified as a substrate. Mutation of Gln356 (Gln233 in E. coli MetAP) to alanine results in a catalytic efficiency about one-third that of native with normal substrates but which can cleave methionine from substrates with penultimate histidine, asparagine, glutamine, leucine, methionine, phenylalanine, and tryptophan. Mutation of Ser195 to alanine had no effect on substrate specificity. None of the altered enzymes produced cleaved substrates with a fully charged residue (lysine, arginine, aspartic acid, or glutamic acid) or tyrosine in the penultimate position.

Aminopeptidases↗
PubMed2001

A common phosphate binding site explains the unique substrate specificity of GSK3 and its inactivation by phosphorylation.

The inhibition of GSK3 is required for the stimulation of glycogen and protein synthesis by insulin and the specification of cell fate during development. Here, we demonstrate that the insulin-induced inhibition of GSK3 and its unique substrate specificity are explained by the existence of a phosphate binding site in which Arg-96 is critical. Thus, mutation of Arg-96 abolishes the phosphorylation of "primed" glycogen synthase as well as inhibition by PKB-mediated phosphorylation of Ser-9. Hence, the phosphorylated N terminus acts as a pseudosubstrate, occupying the same phosphate binding site used by primed substrates. Significantly, this mutation does not affect phosphorylation of "nonprimed" substrates in the Wnt-signaling pathway (Axin and beta-catenin), suggesting new approaches to design more selective GSK3 inhibitors for the treatment of diabetes.

Amino Acid Sequence↗
PubMed1997

The substrate specificity of the snail Lymnaea stagnalis UDP-GlcNAc:GlcNAc beta-R beta 4-N-acetylglucosaminyltransferase reveals a novel variant pathway of complex-type oligosaccharide synthesis.

UDP-GlcNAc:GlcNAc beta-R beta 1-->4-N-acetylglucosaminyltransferase (beta 4-GlcNAcT) of the snail Lymnaea stagnalis is an enzyme with structural resemblance to mammalian beta 4-galactosyltransferases (beta 4-GalT). The enzyme, which is present in the prostate gland of the snail, was expressed in a recombinant form in insect cells using the baculovirus technology. This form was used to determine the enzyme's in vitro substrate specificity in order to assess its possible role in vivo. The enzyme appeared to be a genuine GlcNAcT as no UDP-sugar other than UDP-GlcNAc could serve as an efficient donor substrate. Acceptor specificity studies showed that the enzyme is far more restricted in acceptor usage than beta 4-GalT. Oligomers of beta 4-GlcNAc were relatively poor acceptors, indicating that this enzyme is not involved in the synthesis of chitin-like molecules. Both its polypeptide structure and acceptor specificity suggest that it neither is implicated in the synthesis of the chitobiose core of N-linked glycans. Preferred substrates are those that contain a beta-GlcNAc residue attached to the carbon-6 of Gal or GalNAc residues, as found in vertebrate blood-group I-active and O-linked core 2- and 4-based oligosaccharides, respectively. By contrast, compounds in which GlcNAc is beta 6-linked to Man (as in N-linked glycans) are poor acceptors. Analysis of the products formed in vitro by 1H-NMR spectroscopy and acetolysis showed that the enzyme establishes a GlcNAc beta 1-->4GlcNAc linkage and shows branch specificity with a blood-group I-active trisaccharide substrate. Furthermore, the enzyme differs from beta 4-GalT in that it is not responsive to alpha-lactalbumin. It is proposed that the enzyme functions in a novel, variant pathway of complex-type oligosaccharide synthesis in the snail.

Animals↗
PubMed2003

Characterization of the role of polar amino acid residues within predicted transmembrane helix 17 in determining the substrate specificity of multidrug resistance protein 3.

Human multidrug resistance protein (MRP) 3 is the most closely related homologue of MRP1. Like MRP1, MRP3 confers resistance to etoposide (VP-16) and actively transports 17 beta-estradiol 17-(beta-D-glucuronide) (E(2)17 beta G), cysteinyl leukotriene 4 (LTC(4)), and methotrexate, although with generally lower affinity. Unlike MRP1, MRP3 also transports monovalent bile salts. We have previously demonstrated that hydrogen-bonding residues predicted to be in the inner-leaflet spanning segment of transmembrane (TM) 17 of MRP1 are important for drug resistance and E(2)17 beta G transport. We have now examined the importance of the hydrogen-bonding potential of residues in TM17 of MRP3 on both substrate specificity and overall activity. Mutation S1229A reduced only methotrexate transport. Mutations S1231A and N1241A decreased resistance to VP-16 and transport of E(2)17 beta G and methotrexate but not taurocholate. Mutation Q1235A also reduced resistance to VP-16 and transport of E(2)17beta G but increased taurocholate transport without affecting transport of methotrexate. Mutations Y1232F and S1233A reduced resistance to VP-16 and the transport of all three substrates tested. In contrast, mutation T1237A markedly increased VP-16 resistance and transport of all substrates. On the basis of the substrates analyzed, residues Ser(1229), Ser(1231), Gln(1235), and Asn(1241) play an important role in determining the specificity of MRP3, while mutation of Tyr(1232), Ser(1233), and Thr(1237) affects overall activity. Unlike MRP1, the involvement of polar residues in determining substrate specificity extends throughout the TM helix. Furthermore, elimination of the hydrogen-bonding potential of a single amino acid, Thr(1237), markedly enhanced the ability of the protein to confer drug resistance and to transport all substrates examined.

ATP Binding Cassette Transporter, Subfamily B↗
PubMed1996

Comparison of the substrate specificities of cAMP-dependent protein kinase from bovine heart and Ascaris suum muscle.

The catalytic subunits of cAMP-dependent protein kinases (protein kinase A) from bovine heart and Ascaris suum muscle exhibit only 48% sequence identity and show quantitative differences in substrate specificity. These differences were not obvious at the level of short synthetic substrate peptides but were distinct for some protein substrates. Phosphofructokinase from Ascaris, a physiological substrate, was a better substrate for the protein kinase from the nematode in comparison to the mammalian protein kinase due to a 10-fold lower Michaelis constant. Selective phosphorylation by the two kinases was also observed with some in vitro substrates. In addition, quantitative differences in the interactions between R- and C-subunits from Ascaris and bovine heart were observed. However, several synthetic peptides whose sequence reflected the phosphorylation site of Ascaris suum phosphofructokinase (AKGRSDS*IV), or variations of it, were phosphorylated with the same efficiency by both protein kinases. Based on the data the following are concluded: (1) In agreement with the conservation of structure in the catalytic cleft, the recognition of substrates by protein kinases from phylogenetically distant organisms exhibits similarity. (2) Non-conserved parts of the surface of the protein kinase molecule may contribute to binding of protein substrates and thus to selective recognition.

Amino Acid Sequence↗
PubMed1998

Is one amino acid responsible for substrate specificity of monoamine oxidase A and B?

Mutation of Phe-208 for Ile in monoamine oxidase (MAO) A and mutation of Ile-199 for Phe in MAO B inverted substrate specificity and inhibitor selectivity of the mutants towards the opposite form of the enzyme, MAO B and MAO A, respectively (Y. Tsugeno and A. Ito (1997) J. Biol. Chem., 272, 14033-14036). However, according to data presented by Tsugeno and Ito such point mutations did alter kinetic characteristics of oxidative deamination of some but not all substrates. These point mutations did not alter the secondary structure evaluated by four computer methods. Analysis of the efficacy of a few series of inhibitors suggests spatial differences of substrate binding sites of MAO A and MAO B. All these discrepancies indicate that one amino acid can be responsible for binding of some but not all substrates and inhibitors.

Amino Acid Sequence↗
PubMed1995

Substrate specificity of beta 1,4-N-acetylgalactosaminyltransferase in vitro and in cDNA-transfected cells. GM2/GD2 synthase efficiently generates asialo-GM2 in certain cells.

The substrate specificity of beta 1,4-N-acetylgalactosaminyltransferase has been analyzed using a fusion enzyme which consisted of the catalytic domain of the enzyme and the IgG binding domain of protein A, and also by extracts from cDNA transfectants. Both enzyme sources were capable of producing not only GM2 and GD2, but also asialo-GM2, GalNAc-sialylparagloboside, and Gal-NAc-GD1a from appropriate acceptors, although the efficiencies were at most 1-3% of those of GM2/GD2. The biological significance of these low specificities was studied with transient and stable transfectant cells. From the results of transient expression of the cDNA, asialo-GM2 expression appeared to inversely correlate with GM2 synthase levels in those lines. Consequently, GM2 seemed to be preferentially synthesized when both GM3 and lactosylceramide are available, and asialo-GM2 is synthesized in the absence of GM3 synthesis. However, the results of double immunostaining of CHO transfectants with anti-GM2 and anti-asialo-GM2 antibodies indicated that another factor may be involved in asialo-GM2 synthesis. From the in vitro assay using mixed acceptors, it was concluded that the presence of certain levels of GM2 might enhance the asialo-GM2 synthesis. These results suggest that even acceptors showing low efficiencies in vitro might be used in certain cells depending on the availability of precursors, expression levels of other gangliosides, as well as the kinetic properties of the enzyme, and the compartmentation of the glycosylation machineries in the cells.

Animals↗
PubMed2004

UDP-sugar pyrophosphorylase with broad substrate specificity toward various monosaccharide 1-phosphates from pea sprouts.

UDP-sugars, activated forms of monosaccharides, are synthesized through de novo and salvage pathways and serve as substrates for the synthesis of polysaccharides, glycolipids, and glycoproteins in higher plants. A UDP-sugar pyrophosphorylase, designated PsUSP, was purified about 1,200-fold from pea (Pisum sativum L.) sprouts by conventional chromatography. The apparent molecular mass of the purified PsUSP was 67,000 Da. The enzyme catalyzed the formation of UDP-Glc, UDP-Gal, UDP-glucuronic acid, UDP-l-arabinose, and UDP-xylose from respective monosaccharide 1-phosphates in the presence of UTP as a co-substrate, indicating that the enzyme has broad substrate specificity toward monosaccharide 1-phosphates. Maximum activity of the enzyme occurred at pH 6.5-7.5, and at 45 degrees C in the presence of 2 mm Mg(2+). The apparent K(m) values for Glc 1-phosphate and l-arabinose 1-phosphate were 0.34 and 0.96 mm, respectively. PsUSP cDNA was cloned by reverse transcriptase-PCR. PsUSP appears to encode a protein with a molecular mass of 66,040 Da (600 amino acids) and possesses a uridine-binding site, which has also been found in a human UDP-N-acetylhexosamine pyrophosphorylase. Phylogenetic analysis revealed that PsUSP can be categorized in a group together with homologues from Arabidopsis and rice, which is distinct from the UDP-Glc and UDP-N-acetylhexosamine pyrophosphorylase groups. Recombinant PsUSP expressed in Escherichia coli catalyzed the formation of UDP-sugars from monosaccharide 1-phosphates and UTP with efficiency similar to that of the native enzyme. These results indicate that the enzyme is a novel type of UDP-sugar pyrophosphorylase, which catalyzes the formation of various UDP-sugars at the end of salvage pathways in higher plants.

Amino Acid Sequence↗
PubMed1994

Investigation of the substrate specificity of a cloned expressed human bilirubin UDP-glucuronosyltransferase: UDP-sugar specificity and involvement in steroid and xenobiotic glucuronidation.

A cloned human bilirubin UDP-glucuronosyltransferase (UGT) stably expressed in Chinese hamster V79 cells was used to assess the substrate specificity of the enzyme. The catalytic potential (Vmax/Km(bilirubin) of the enzyme with UDP-glucuronic acid (UDPGA) was 2-fold and 10-fold greater than that for UDP-xylose and UDP-glucose respectively. The formation of bilirubin mono- and di-conjugates was found to be dependent on time, UDP-sugar concentration and bilirubin concentration. Ex vivo studies demonstrated that the genetically engineered cell line was capable of the uptake and glucuronidation of bilirubin and the release of bilirubin glucuronide, indicating its usefulness in studying transport processes. Over 100 compounds, including drugs, xenobiotics and endogenous steroids, were tested as substrates for the enzyme to determine the chemical structures accepted as substrates. A wide diversity of xenobiotic compounds such as phenols, anthraquinones and flavones (many of which are in foodstuffs) were glucuronidated by the enzyme. The enzyme also had the capacity to glucuronidate oestriols and oestradiols stereoselectively. H.p.l.c. analysis of the regioselective glucuronidation of beta-oestradiol (E2) demonstrated that it was conjugated solely at its A-ring hydroxy group by the bilirubin UGT to form E2-3-glucuronide, this was in contrast with human liver microsomes which formed 3- and 17-glucuronides of this oestrogen. Studies utilizing microsomes from a Crigler-Najjar patient and inhibition of E2 glucuronidation with bilirubin indicated that the cloned expressed bilirubin UGT was the major human UGT isoform responsible for the formation of E2-3-glucuronide, which is the predominant E2 conjugate in human urine.

Animals↗
PubMed2002

pH, inhibitor, and substrate specificity studies on Escherichia coli penicillin-binding protein 5.

The recent structural determination of Escherichia coli penicillin-binding protein 5 (PBP 5) provides the opportunity for detailed structure-function studies of this enzyme. PBP 5 was investigated in terms of its stability, linear reaction kinetics, acyl-donor substrate specificity, inhibition by a number of active site-directed reagents, and pH profile. PBP 5 demonstrated linear reaction kinetics for up to several hours. Dilution of PBP 5 generally resulted in substantial loss of activity, unless BSA or a BSA derivative was added to the diluting buffer. PBP 5 did not demonstrate a significant preference against a simple set of five alpha- and epsilon-substituted L-Lys-D-Ala-D-Ala derivatives, suggesting that PBP 5 lacks specificity for the cross-linked state of cell wall substrates. Among a number of active site-directed reagents, only some thiol-directed reagents gave substantial inhibition. Notably, serine-directed reagents, organic phosphates, and simple boronic acids were ineffective as inhibitors. PBP 5 was stable over the pH range 4.6-12.3, and the k(cat)/K(m) vs. pH profile for activity against Ac(2)-L-Lys-D-Ala-D-Ala was bell-shaped, with pK(a)s at 8.2 and 11.1. This is the first complete pH profile, including both acidic and basic limbs, for a PBP-catalyzed DD-carboxypeptidase (CPase) reaction. Based on its structure, similarity to Class A beta-lactamases, and results from mutagenesis studies, the acidic and basic limbs of the pH profile of PBP 5 are assigned to Lys-47 and Lys-213, respectively. This assignment supports a role for Lys-47 as the general base for acylation and deacylation reactions.

Animals↗
PubMed2002

Signal sequences control gating of the protein translocation channel in a substrate-specific manner.

N-terminal signal sequences mediate targeting of nascent chains to the endoplasmic reticulum and facilitate opening of the protein translocation channel to the passage of substrate. We have assessed each of these steps for a diverse set of mammalian signals. While minimal differences were seen in their targeting function, signal sequences displayed a remarkable degree of variation in initiating nascent chain access to the lumenal environment. Such substrate-specific properties of signals were evolutionarily conserved, functionally matched to their respective mature domains, and important for the proper biogenesis of some proteins. Thus, the sequence variations of signals do not simply represent functional degeneracy, but instead encode critical differences in translocon gating that are coordinated with their respective passengers to facilitate efficient translocation.

3T3 Cells↗
PubMed1999

Substrate specificity of a nitroalkane-oxidizing enzyme.

The flavoprotein nitroalkane oxidase from Fusarium oxysporum catalyzes the oxidation of nitroalkanes to aldehydes with production of hydrogen peroxide and nitrite. The substrate specificity of the FAD-containing enzyme has been determined as a probe of the active site structure. Nitroalkane oxidase is active on primary and secondary nitroalkanes, with a marked preference for unbranched primary nitroalkanes. The V/K values for primary nitroalkanes increase with increasing length of the alkyl chain, reaching a maximum with 1-nitrobutane, suggesting a hydrophobic binding site sufficient to accommodate a four carbon chain. Each methylene group of the substrate contributes approximately 2.6 kcal mol-1 in binding energy. The V/K values for substrates containing a hydroxyl group are two orders of magnitude smaller than those of the corresponding nitroalkanes, also consistent with a hydrophobic binding site. 3-Nitro-1-propionate is a competitive inhibitor with a Kis value of 3.1 +/- 0.2 mM.

Aldehydes↗
PubMed2001

Recombinant tyrosine aminotransferase from Trypanosoma cruzi: structural characterization and site directed mutagenesis of a broad substrate specificity enzyme.

The gene encoding tyrosine aminotransferase (TAT, EC 2.6.1.5) from the parasitic protozoan Trypanosoma cruzi was amplified from genomic DNA, cloned into the pET24a expression vector and functionally expressed as a C-terminally His-tagged protein in Escherichia coli BL21(DE3)pLysS. Purified recombinant TAT exhibited identical electrophoretic and enzymatic properties as the authentic enzyme from T. cruzi. Both recombinant and authentic T. cruzi TATs were highly resistant to limited tryptic cleavage and contained no disulfide bonds. Comprehensive analysis of its substrate specificity demonstrated TAT to be a broad substrate aminotransferase, with leucine, methionine as well as tyrosine, phenylalanine, tryptophan and alanine being utilized efficiently as amino donors. Valine, isoleucine and dicarboxylic amino acids served as poor substrates while polar aliphatic amino acids could not be transaminated. TAT also accepted several 2-oxoacids, including 2-oxoisocaproate and 2-oxomethiobutyrate, in addition to pyruvate, oxaloacetate and 2-oxoglutarate. The functionality of the expression system was confirmed by constructing two variants; one (Arg389) being a completely inactive enzyme; the other (Arg283) retaining its full activity, as predicted from the recently solved three-dimensional structure of T. cruzi TAT. Thus, only one of the two strictly conserved arginines which are essential for the enzymatic activity of subfamily Ialpha aspartate and aromatic aminotransferases is critical for T. cruzi's TAT activity.

Amino Acids↗
PubMed1979

The substrate specificity of yeast hexokinase: reaction with D-arabinose oxime.

By chromatography, electrophoresis, n.m.r. spectroscopy, and spectrophotometric assay, it has been shown that D-arabinose oxime acts as a weak substrate for yeast hexokinase. The enzyme-catalysed phosphorylation of the oxime, which exists as a mixture of E (80%) and Z (20%) acyclic forms in solution at equilibrium, is proposed to proceed via the transient formation of a furanoid species. Weak substrate-activity was also observed with 4-deoxy-D-xylo-hexose, but not with 5-deoxy-D-xylohexose. The relation of these and previous results concerning the carbohydrate-substrate specificity of yeast hexokinase in solution to X-ray crystallographic studies is discussed.

Arabinose↗
PubMed1981

Substrate specificity of lysophospholipase-transacylase from rat lung and its action on various physical forms of lysophosphatidylcholine.

Lysophospholipase-transacylase (lysolecithin acylhydrolase, EC 3.1.1.5) from rat lung catalyzes the transfer of acyl groups from lysophosphatidylcholine to either water or another molecule of lysophosphatidylcholine. Studies on the substrate specificity of the purified enzyme showed that a phosphate group in the substrate is essential for enzymatic activity; monoacylglycerol is not hydrolyzed, nor does it serve as an acceptor of acyl groups. The influence of the acyl chain in lysophosphatidylcholine was investigated by using mixtures of differently labelled lysophosphatidylcholine species, or by studying the transfer of [1-14C]Palmitate from [1-14C]palmitoylpropane (1,3)diol-phosphocholine to various 1-acyl-sn-glycero-3-phosphocholines. Lysophosphatidylcholines with acyl chains comprised of ten or more C-atoms were found to serve as acyl acceptors. This finding was used to determine the action of the enzyme on 1-[1-14C]auroyl- and 1[1-14C]myristoyl-sn-glycero-3-phosphocholine both below and above the critical micelle concentration of the substrate. Monomeric substrate was effectively hydrolyzed, but the transacylase activity of the enzyme was only expressed when substrate micelles were present. Likewise, no transacylase activity was found when lysophosphatidylcholine was embedded in liposomal membranes prepared from lung total lipids. These findings, which persist with crude enzyme preparations (100 000 x g supernatant), are discussed in relation to the putative function of the lysophospholipase-transacylase in the synthesis of disaturated phosphatidylcholine in lung.

Acyltransferases↗