Restoration Of Enzyme Activity Research Articles

Regulation of liver phosphorylase phosphatase by glutathione disulfide.

The mechanism of inactivation of rabbit liver phosphorylase phosphatase by glutathione disulfide (GSSG) was investigated. The catalytic subunit of phosphorylase phosphatase was inactivated by GSSG and other disulfides. Inactivation by GSSG was a concentration-dependent process and resulted in the formation of an inactive, stable enzyme species. The inactivated enzyme could be reactivated by addition of various sulfhydryl compounds, including glutathione (GSH). Homogeneous phosphorylase phosphatase contains, per mol of catalytic subunit (Mr = 33,000), two sulfhydryl groups, one of which reacted with GSSG to form inactive enzyme. Binding studies with [glycine-2-3H]GSSG revealed simultaneous incorporation of 3H radioactivity into the catalytic subunit and stoichiometric loss of catalytic activity. Treatment of the 3H-labeled enzyme with GSH was accompanied by release of 3H radioactivity from the enzyme and restoration of enzyme activity. The results suggest that inactivation of phosphorylase phosphatase by GSSG results from the formation of a mixed disulfide between GSSG and one of the two sulfhydryl groups in the catalytic subunit.

Mechanism of porphobilinogen synthase. Requirement of Zn2+ for enzyme activity.

The role of metal ions in the mechanism of action of bovine liver porphobilinogen synthase was investigated. Studies with chelating agents were consistent with a requirement of metal ions for enzyme activity, and the use of 8-hydroxyquinoline-5-sulfonic acid suggested that Zn2+ was present in the enzyme. The low activity detected in metal-free apoporphobilinogen synthase was attributed to adventitious metal ions. Addition of Zn2+ to the apoenzyme completely restored enzyme activity if the essential sulfhydryl groups on the enzyme were first reduced with sulfhydryl reagents. It does not follow necessarily from this observation that Zn2+ forms a bond with a sulfhydryl group in the enzyme. However, we also observed that Zn2+ did not bind to the enzyme unless the essential cysteinyl residues were reduced. We have concluded that the octameric enzyme contains 4 g atoms of Zn2+/mol from our enzyme activity measurements and binding studies. Alkylation of the enzyme resulted in a marked reduction in the binding of Zn2+ to the enzyme. These observations are consistent with the suggestion that the interaction of the Zn2+ ions with the enzyme occurs with sulfhydryl groups at the active site. It appears that Zn2+ does not participate in substrate binding nor in the maintenance of the quaternary structure of the enzyme. Possible mechanistic roles for Zn2+ in porphobilinogen synthase are discussed. It should be noted that Cd2+ was the only other element found which restored activity to the apoenzyme.

A new class of monoamine oxidase inhibitors.

Abstract:Newly synthesized compounds have been found to inhibit mitochondrial monoamine oxidase (MAO) in mouse brain and rat liver. A series of 2‐acylamino‐3‐tert‐aminopropiophenones acted preferentially against MAO type B (2‐phenylethylamine as substrate), apparently irreversibly. 2‐Decanoylamino‐3‐morpholinopropiophenone acted similarly in vivo toward the cerebral MAO, producing a dose‐related inhibition. At high dose levels, MAO type A was also severely inhibited. The effects were produced rapidly and restoration of enzyme activity also appeared rapidly. The half‐life for MAO type A could be estimated from the rate of enzyme reappearance to be 13 h. It is suggested that the amino ketones undergo a β‐elimination reaction at the enzyme's active site, forming a reactive species (an α,β‐unsaturated ketone), which reacts covalently with a nucleophilic group of the enzyme by a Michael addition. Some other related compounds, derivatives of phenylpropane, also showed inhibitory activity against MAO, particularly against type A (serotonin as substrate). The morpholino compound might have promise as a quickly effective, short‐acting inhibitor of MAO type B.

Electrophoretic analysis of plasminogen activators in polyacrylamide gels containing sodium dodecyl sulfate and copolymerized substrates

A new technique is described for the electrophoretic analysis of plasminogen activators in sodium dodecyl sulfate-polyacrylamide gels containing copolymerized plasminogen and gelatin. The method depends upon the fact that the zymogen and gelatin, when incorporated into the polyacrylamide matrix at the time of casting, are retained during subsequent electrophoresis of enzyme samples, and serve as satisfactory sequential, in situ substrates for the localization of plasminogen activator bands by negative staining. The nonionic detergent, Triton X-100, is used to remove sodium dodecyl sulfate and restore enzyme activity. The method can be used to detect as little as 1 mU of urokinase and effectively distinguishes between melanoma- and urokinase-type plasminogen activators. Plasminogen-independent proteases are detected by omission of plasminogen from the gel.

Molecular basis for genetic complementation in propionyl CoA carboxylase deficiency

The extent of genetic complementation in polyethylene glycol-induced heterokaryons of propionyl CoA carboxylase deficient fibroblast lines was determined by comparing enzyme activity changes over time in pairwise fusions of the three major complementation groups, bio, pcc A and pcc C, with the activity changes in similarly mixed but unfused cultures. Maximum complementation between bio and pcc. A or pcc C lines was attained within 24 h after fusion and was not inhibited by cycloheximide. In contrast, the complementation between pcc A and pcc C lines only attained 50% of the maximum restored carboxylase activity by 24–36 h and the increase was 93% inhibited by cycloheximide. Maximum restoration of activity was not achieved until 72–96 h after fusion. Removal of cycloheximide at 24 h permitted complementation to take place. Our studies suggest that intergenic complementation between the bio and pcc A or pcc C lines is due to the contribution within the heterokaryon of normal enzymes from each of the respective lines, resulting in almost immediate, protein synthesis-independent, partial restoration of carboxylase activity. Complementation between pcc A and pcc C lines also appears to be intergenic but probably results from the de novo synthesis of normal subunits and protomers which assemble into normal or stabilized propionyl CoA carboxylase molecules with restored enzyme activity.

5'-p-fluorosulfonylbenzoyladenosine as an ATP site affinity probe for Na+, K+-ATPase.

We have investigated the suitability of 5'-p-fluorosulfonylbenzoyladenosine (FSBA) as an ATP site affinity probe for the canine kidney Na+, K+-ATPase. The purified enzyme is slowly inactivated by this compound in suitable buffers, losing about half of its activity over a two-hour period. The rate of inactivation is more rapid in 0.1 M KCl than in 0.1 M NaCl. Low concentrations of ATP protect the enzyme against inactivation, with half-maximal effects at 4 microM ATP in 0.1 M NaCl and 350 microM ATP in 0.1 M KCl. ADP also protects against FSBA inhibition, but AMP is ineffective when present at 100 microM levels. This pattern is consistent with the previously described nucleotide specificity of the Na+, K+-ATPase. Addition of protective amounts of ATP after inactivation has occurred does not restore enzyme activity, indicating that inhibition is irreversible. Measurement of the concentration-dependence of FSBA inactivation suggests an apparent Kd for binding of this compound well above 1 mM, the solubility limit of the analog. This finding is reinforced by the failure of 1 mM FSBA to compete effectively with ATP for the high-affinity ATP site of the enzyme. Nevertheless, attachment of the analog to this site is indicated by its ability to prevent [3H]-ADP binding in proportion to the number of sites it has inactivated. Studies with [3H]-FSBA show that about 1 mole of the analog attaches specifically to the alpha subunit per mole of enzyme inactivated. A similar amount of nonspecific labeling also occurs with negligible effect on enzyme activity. These findings suggest that FSBA may be useful in probing the topography of the high-affinity ATP binding site of the Na+, K+-ATPase and related enzymes.

Chromosomal assignment of the gene for folylpolyglutamate synthetase to human chromosome 9.

An auxotrophic mutant, GAT-, derived from the Chinese hamster cell line CHO-K1 and exhibiting multiple growth requirements for glycine, adenine, and thymidine, has been shown to be deficient in one of the folate-dependent enzymes, folylpolyglutamate synthetase (FPGS). This mutant was fused with normal human lymphocytes and human lymphoblasts and 41 GAT+ primary hybrid clones were isolated in medium lacking glycine, adenine, and thymidine. In addition, 71 secondary clones were isolated after growth of four primary hybrid clones in enriched medium, which allows losing the complementing human chromosome without losing cell viability. Analysis of human isozyme markers in the 112 primary and secondary clones established a syntenic relationship between the GAT marker and AK1, an isozyme marker for human chromosome 9. Enzyme studies of the parental and hybrid cells not only showed restored enzyme activities of FPGS in the hybrids, but also demonstrated that several enzyme characteristics of FPGS in the hybrids are consistent with those of the human enzyme. Thus, it is concluded that the human gene coding for the enzyme FPGS which complements the auxotrophic mutant GAT- in the CHO-K1 cells can be assigned to human chromosome 9. This assignment provides an additional selective marker for mammalian cell genetic analysis in which human chromosome 9 can be selectively and stably retained in the cell hybrids.

Evidence for an essential histidyl residue at the active site of pyridoxamine (pyridoxine)-5'-phosphate oxidase from rabbit liver.

Pyridoxamine (pyridoxine)-5'-phosphate oxidase (EC 1.4.3.5) from rabbit liver is inactivated by diethylpyrocarbonate in an all-or-none fashion with first order kinetics with respect to modifier concentration. The rate of inactivation increases with pH and reflects a group with a pKa of 7.5. Inactivated enzyme is in the holo form with intact FMN. Four histidyls and a cysteinyl residue are modified by excess reagent. The restoration of enzymatic activity by hydroxylamine, the spectrophotometric and colorimetric amino acid analyses, and our previous studies on cysteine modification (Tsuge, H., and McCormick, D.B. (1979) in Flavins and Flavoproteins (Yamano, T., and Yagi, K., eds) Japan Scientific Societies Press, Tokyo, in press) all suggest that inactivation occurs solely by modification of histidine. Analyses by kinetic and statistical methods indicate that three histidines are modified slowly and are not critical for activity, while one histidine is modified nine times more rapidly and accounts for the observed inactivation. Inactivated enzyme shows no significant perturbations in structure, as evidenced by absorption, CD, fluorescence, and gel filtration, but is unable to bind the product, pyridoxal 5'-phosphate. Furthermore, the substrate-competitive inhibitor, pyridoxal 5'-phosphate oxime, protects from inactivation. Hence, diethylpyrocarbonate inactivates this enzyme by modifying a crucial histidyl residue at the substrate/product-binding site.

Serine hydroxymethylase. Specificity of bond cleavage to form quinonoid intermediates and rate of holoenzyme formation

l-Serine transhydroxymethylase (5,10-methylenetetrahydrofolate:glycine hydroxymethyltransferase, EC 2.1.2.1) a pyridoxal phosphate-dependent enzyme, has been obtained as a homogeneous preparation with a specific activity of 6.7 μmol benzaldehyde per minute at 30°C at pH 7.5 in N-2- hydroxyethylpiperazine-N′-2- ethanesulfonic acid (Hepes) buffer, with dl-threo-β-phenylserine as a substrate. This enzyme has been used to study the specificity of bond cleavage in forming quinonoid intermediates from dl and non-asymmetric amino acids. The ability of the generated quinonoids to react with formaldehyde and acetaldehyde has also been studied and evidence obtained for formation of the corresponding β-hydroxymethyl and β-hydroxyethyl amino acid derivaties. Apotranshydroxymethylase has been prepared and the rate of holoenzyme formation was found to be 0.52 min −1 by measuring Schiff base formation at 425 nm and 0.66 min −1 as determined from restoration of enzymic activity. A requirement for the presence of mercaptoethanol for complete reactivation was also established by these studies.

A dimer--dimer binding region in beta-galactosidase.

alpha Complementation in beta-galactosidase is the restoration of enzyme activity by addition of the alpha donor CNBr2, from amino acid residues 3--92 of the polypeptide, to inactive M15 protein from the lacZ deletion mutant strain M15. M15 protein lacks residues 11--41 and is a dimer; the active complex, like native beta-galactosidase, is tetrameric [Langley, K. E., & Zabin, I. (1976) Biochemistry 15, 4866--4875]. A dimer--dimer binding region in beta-galactosidase has been identified by proteolytic and immunologic studies of alpha-complementation. Proteolytic experiments were carried out with trypsin. Treatment of native beta-galactosidase with trypsin, followed by reaction of the mixture with cyanogen bromide, yields intact CNBr2 as measured by its ability to complement M15 protein. Active CNBr2 is not obtained when urea-denatured beta-galactosidase is treated in the same way. Therefore the segment corresponding to CNBr2 is apparently buried within the folded protein. Immunologic experiments were carried out with antibodies against CNBr2, tryptic peptide T8 (residues 60--140), and CNBr3 (residues 93--187). Anti-CNBr2 and anti-T8 bind to M15 protein but not to beta-galactosidase, indicating that this area is exposed in the dimer. Anti CNBr2, but not anti-T8 or anti-CNBr3, inhibits the formation of alpha-complemented enzyme. These results indicate that an early part of the sequence, within the segment corresponding to CNBr2, is involved in dimer--dimer interaction.

Lipoprotein lipase of cultured mesenchymal rat heart cells: III. Effect of glucocorticoids and insulin on enzyme formation

Lipoprotein lipase activity was studied in mesenchymal cells isolated from rat hearts and cultured for up to 8 days. The enzyme activity increased markedly between day 3 and 5 while the subsequent increase was less pronounced. Addition of hydrocortisone to complete culture medium resulted in an increase in lipoprotein lipase activity at all stages of culture. Lipoprotein lipase activity did not increase after addition of insulin to the complete culture medium. In the presence of serum-poor medium between day 3 and 6, the increase in lipoprotein lipase activity was much lower than in the presence of complete culture medium. Addition of hydrocortisone and insulin to the serum-poor medium resulted in a significant rise in lipoprotein lipase activity while less consistent effects were obtained after addition of each hormone alone. Transfer of cells to serum-poor medium between day 6 and 7 of culture caused a fall in enzyme activity. Addition of hydrocortisone alone and with insulin restored enzyme activity to control values. No effect on lipoprotein lipase was seen with estradiol, growth hormone, or glucagon when added to serum-containing medium, or serum-poor medium. These results indicate that the lipoprotein lipase of heart is controlled by glucocorticoids and that this control might require the presence of insulin for optimal expression.

THE NATURE OF GENETIC INSTABILITY IN AUXOTROPHS OF SALMONELLA TYPHIMURIUM REQUIRING CYSTEINE OR METHIONINE AND RESISTANT TO INHIBITION BY 1,2,4-TRIAZOLE

ABSTRACT We tested the hypothesis that unstable suppression of auxotrophy in triazole-resistant derivatives of Cym- mutants of Salmonella typhimurium is due to reversible insertion at the Cym- site of genetic material originating in the cysALKptsHI region. We have shown that the unstable phenotype was co-transducible with markers in the cysCDHIJ region. The suppression of the Cym phenotype was recA dependent and frequencies of segregation were affected by UV irradiation. Restored enzyme activity in suppressed strains was determined by wild-type enzyme, suggesting that the unstable regions are located in cys gene regulatory regions. These results support the hypothesis. In contradiction, we found no evidence for a deletion in the cysALKptsHI region.

Restoration of enzyme activity by recessive missense suppressors in the fungus Coprinus.

The acu-1 locus in Coprinus is the structural gene for acetyl-CoA synthetase. Five suppressor gene mutations, which suppress the acu-1,34 missense allele, were induced by mutagen treatment. All five suppressors were shown to have properties expected for tRNA structural gene mutations: they are recessive, they show a gene dosage effect in any doubly heterozygous combination of two sup+ mutations and they are allele specific in action.

Site of ATPase activity in Myxococcus xanthus: lipid requirement for enzyme activity. Dedicated to Professor Dr. W. Schwartz on his 80th birthday.

Treatment of cells with lysophosphatidylcholine, lysozyme, and phospholipase D removed most of their phospholipids and reduced ATPase activity to near zero. Addition of a microdispersion of phospholipids restored enzyme activity to various degrees. Phosphatidylcholine was most effective in reconstitution experiments, less effective were phosphatidylethanolamine and phosphatidylserine. Lipid analyses of cell fractions were possible through separation of cell wall and cell membrane in a sucrose gradient after differentiated treatment of glutaraldehyde fixed cells with lysophosphatidylcholine, lysozyme, and pronase. Phosphatidylcholine was almost exclusively a component of the cell membrane, whereas phosphatidylethanolamine was that of the wall. It is concluded that lipids are necessary for in vivo function of a Mg-dependent ATPase, and that membrane-associated phosphatidylcholine may serve as a matrix for the enzyme. Lipid extracts made from cells or cell fractions contained plasmologens, not previously reported to occur in Gram-negative, aerobic bacteria.

Site of ATPase activity in Myxococcus xanthus: Lipid requirement for enzyme activity

AbstractTreatment of cells with lysophosphatidylcholine, lysozyme, and phospholipase D removed most of their phospholipids and reduced ATPase activity to near zero. Addition of a microdispersion of phospholipids restored enzyme activity to various degrees. Phosphatidylcholine was most effective in reconstitution experiments, less effective were phosphatidylethanolamine and phosphatidylserine. Lipid analyses of cell fractions were possible through separation of cell wall and cell membrane in a sucrose gradient after differentiated treatment of glutaraldehyde fixed cells with lysophosphatidylcholine, lysozyme, and pronase. Phosphatidylcholine was almost exclusively a component of the cell membrane, whereas phosphatidylethanolamine was that of the wall. It is concluded that lipids are necessary for in vivo function of a Mg‐dependent ATPase, and that membrane‐associated phosphatidylcholine may serve as a matrix for the enzyme. Lipid extracts made from cells or cell fractions contained plasmologens, not previously reported to occur in Gram‐negative, aerobic bacteria.

Restoration of β-galactosidase to Escherichia coli M15. Complementation studies

Carboxymethylated beta-galactosidase from Escherichia coli was dissociated at 100 degrees C to form carboxymethylated fragments A and B. The mol.wts. of carboxymethylated fragments A and B were determined by gel filtration to be 64300 and 22400 respectively. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis of carboxymethylated fragments A and B that had been pretreated with 2-mercaptoethanol and sodium dodecyl sulphate yielded mol.wts. of 64000 and 22100 respectively. Carboxymethylated fragments A and B had arginine as their C-terminal amino acid. When a crude extract of E. coli M15 was filtered through a column of Sepharose 6B, it was found that carboxymethylated fragment B could restore beta-galactosidase activity when added to fractions having mol.wts. estimated to be 123000, 262000 and 506000. These fractions are referred to as ;complementable fractions'. Similarly, it was found that carboxymethylated fragment A could restore enzyme activity to tractions having mol.wts. estimated to be 63000, 253000 and 506000. Estimates of the molecular weights of the beta-galactosidase activity obtained by restoration with carboxymethylated fragments A and B were made by filtering the active enzyme through another column of Sepharose 6B. The enzyme obtained by complementation with carboxymethylated fragment B, i.e. the complemented enzyme, had mol.wt. 525000, and that obtained with carboxymethylated fragment A had mol.wts. of 525000, 646000 and 2000000. The latter finding suggests that multiple forms of complemented beta-galactosidase can exist.

Effects of lipids on the activity of ferrochelatase

Removal of lipids from submitochondrial particles or detergent-solubilized mitochondrial preparations of rat liver resulted in a 90% loss of ferrochelatase (protochemeferro-lyase, EC 4.99.1.1) activity. The addition of either a fatty acid or phospholipid restored enzyme activity; the extent of reactivation being correlated with the degree of unsaturation of the fatty acid or acyl chain and independent of the polar head group of the phospholipid. Arrhenius plots of the ferrochelatase activities of submitochondrial particles and detergent-solubilized mitochondrial preparations showed transition temperatures of 37 and 28.5°C, respectively. Ferrochelatase of submitochondrial particles or detergent-solubilized preparations had an absolute requirement for Ca 2+. The ferrous salt of oxalic acid, a Ca 2+ chelator, was a very poor substrate for these preparations. In contrast, ferrochelatase activities of fatty acid- or lipid supplemented acetone extracts of these preparations were not dependent on the presence of Ca 2+ and ferrous oxalate served as substrate for these extracts.

Studies on the Restoration of the Activities of Ribonucleases by Polyamines in the Presence of Various Ribonuclease Inhibitors

The effect of polyamines on ribonucleases in the presence of various inhibitors (poly(G), heparin, and rat liver RNase inhibitor) has been studied. Bovine pancreatic RNas A and a ribonuclease from horse submaxillary gland (RNase HS) were inhibited by the inhibitors, but RNase T1 and RNase M were not inhibited. Polyamines were found to restore the activites of RNase A and RNase HS inhibited by poly(G) or heparin but not those activities inhibited by rat liver RNase inhibitor. When poly(U) and poly(C) were used as substrates, the inhibitory effects of poly(G) and heparin were greater with poly(U) than poly(C) as a substrate. However, when poly(C) was used as a substrate in the presence of either of the above inhibitors, the restoration of RNase activity by sperimine was more efficient. In fact, a stimulatory effect was observed. From the double-reciprocal plots, it was concluded that polyamines restored the activiities of RNases by increasing the availability of the substrate and enzyme to each other. The restoration of enzyme activity by polyamines occurred through the binding of the polyamines to the inhibitor and the subsequent release of enzyme from the inhibitor.

Interaction of rifamycins with mammalian nucleic acid polymerizing enzymes

Procedures were established for the isolation and partial purification of DNA polymerase, RNA polymerase and poly(A) polymerase activities from the cytoplasm and nuclei of NIH-Swiss mouse embryos. Based on the elution pattern of these enzyme activities from DEAE-cellulose and phosphocellulose columns in Tris · HCl buffer, pH 8.0, the apparent basicities of the enzymes can be arranged as follows: cytoplasmic(C) poly(A) polymerase > (C)DNA polymerase β > (C)DNA polymerase α and nuclear(N) poly(A) polymerase > (N)DNA polymerase > (N)RNA polymerase I > (N)RNA polymerase II. Twenty rifamycins, including rifamycin B, rifamycin S, rifamycin SV, and rifamycin SV derivatives, were examined for their ability to inhibit the above mentioned nucleic acid polymerizing enzymes and Simian sarcoma virus type I (SSV-1) reverse transcriptase. Rifamycin SV 3′-formyldiphenylhydrazone, rifamycin SV 3′-formyl-n-octyloxime (AF/013) and rifamycin SV 3′-formyldiphenylmethyloxime (AF/05) inhibited all the tested enzyme activities. Rifamycin SV 3′-formylpropylphenyloxime (AF/015) inhibited cellular nucleic acid polymerase activities but not SSV-1 DNA polymerase activity. Rifamycin SV 3′-formyldinitrophenylhydrazone (AF/DNFI) strongly inhibited reverse transcriptase activity but did not inhibit cellular DNA polymerase activities. AF/DNFI slightly inhibited RNA and poly(A) polymerase activities. Rifamycin SV 3′-formyldipropylhydrazone (AF/DPI) and 2,6-dimethyl-4- N-benzyldemethylrifampicin (AF/ABDMP) slightly inhibited reverse transcriptase activity but did not inhibit cellular nucleic acid polymerase activities. Active rifamycin derivatives inhibited enzyme reactions by interacting with the enzyme proteins. Nascent polynucleotide chain elongation continued although at a reduced rate in the presence of inhibitor. The addition of increasing concentrations of nonionic detergent (Triton X-100) to rifamycin-inhibited enzyme reactions fully restored enzyme activities. The presence of highly lipophilic 3′-side chains on active rifamycins and the reversibility of enzyme inhibition by Triton X-100 suggest that the tested nucleic acid polymerizing enzymes may have hydrophobic regions with which inhibitory rifamycins interact.

Interaction of concanavalin a with membrane-bound and solubilized lipoprotein lipase of rat heart

Concanavalin A was used to study the configuration of lipoprotein lipase at the surface of capillary endothelium. Incubation of heart homogenates with increasing concentrations of concanavalin A for 5–60 min resulted in inhibition of up to 50% of enzyme activity. The inhibition was related to the concentration of lectin and the time of incubation and was fully reversible by postincubation with α-methyl- d-mannoside. Rat hearts were perfused for 5–60 min and lipoprotein lipase activity determined in postheparin perfusates and in the perfused heart. When the lectin was introduced into the perfusate a significant reduction of heparin-releasable enzyme was found after 30 min of perfusion. The missing enzyme could be recovered by postperfusion with α-methyl- d-mannoside, but not by addition of the sugar to the perfusate withdrawn from the apparatus. These results suggested binding of lectin to the surface-located enzyme and support for such a binding was obtained by the finding of release of labeled lectin into the perfusate by heparin. Perfusion of hearts with concanavalin A for 60 min resulted also in a fall in nonreleasable lipoprotein lipase. The mechanism of this fall is not due to impairment of enzyme synthesis, as leucine incorporation into protein was not reduced. Since neither perfusion nor postincubation with α-methyl- d-mannoside restored enzyme activity, the fall was most probably due to irreversible inhibition. It is concluded that mannose residues of lipoprotein lipase in heart homogenates and at the endothelial surface of heart capillaries are available to interact with a specific lectin. Such an interaction renders the enzyme less releasable by heparin during perfusion and causes a significant inhibition of enzyme activity in homogenates.