Characterization of human growth hormone from acromegalic pituitary tumors

Human growth hormone (HGH) was extracted from acromegalic pituitary tumors at pH 10.5 and precipitated with ammonium sulfate at 20-40% saturation. It was purified on a Sephadex G-100 column to yield monomeric HGH. The tumor-HGH was indistinguishable from the authentic one in polyacrylamide gel electrophoresis at pH 8.3 or in the presence of sodium dodecyl sulfate, high-performance liquid chromatography, radioimmunoassay, peptide map, amino acid composition and N-terminal partial amino acid sequence. The tumor-HGH is active in the tibia assay and bodyweight gain test in hypophysectomized rats with comparable potency to that of the authentic sample.     

Purification and properties of two ribonucleases and a nuclease from barley seeds

Three enzymes possessing RNAase activity were isolated from barley seeds. These enzymes were further purified by ammonium sulphate precipitation DEAE-cellulose chromatography, gel filtration on Sephadex G-75 and DEAE-Sephadex A-50 chromatography. These enzymes have been characterized and classified as: 1. Plant RNAase I (EC 3.1.27.1). It has a pH optimum at 5.7 and molecular weight of 19 000. 2. Plant RNAase II (EC 3.1.27.1). It has a pH optimum at 6.35 and molecular weight of 19 000. 3. Plant nuclease I (EC 3.1.30.2). It has a pH optimum at 6.8 and molecular weight of 37 000. Two RNAases were purified to homogeneity by means of affinity chromatography on poly(G)-Sepharose 4B, as shown by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate.     

Structural and chemical characterization of a homogeneous peptide N-glycosidase from almond

A peptide N-glycosidase that catalyzes the hydrolysis of N-linked oligosaccharide chains from glycopeptides and glycoproteins has been purified to homogeneity from almond emulsin and from almond meal. Purification from almond emulsin using ion-exchange chromatography, gel filtration chromatography, and preparative polyacrylamide gel electrophoresis gave an enzyme which was purified more than 700-fold and with a yield of 63%. An alternative procedure, more suitable for efficient large scale purification, used ion-exchange, affinity, and gel filtration chromatography. When purification began with almond emulsin, the enzyme was purified 1200-fold with a 37% yield, while when purification began with almond powder, the enzyme was purified 9000-fold with a yield of 45%. The homogeneous enzyme is stable at 4 degrees C for several months in 10 mM sodium acetate, pH 5.0, buffer. The peptide N-glycosidase is itself shown to be a glycoprotein consisting of a single polypeptide chain with a molecular weight of 66 800 on sodium dodecyl sulfate (SDS)-polyacrylamide gel electrophoresis. Circular dichroism spectra of the native molecule indicate the presence of a high (approximately 80%) alpha-helix content. The amino acid and carbohydrate contents of the enzyme are presented. When a convenient new assay with a turkey ovomucoid glycopeptide as a substrate is used, the enzyme preparation exhibits a broad pH optimum centered between pH 4 and pH 6. The enzyme is readily inactivated by SDS and guanidine hydrochloride, but it is stable in the presence of moderate concentrations of several other protein denaturants.(ABSTRACT TRUNCATED AT 250 WORDS)     

Purification and characterization of acid trehalase from the yeast suc2 mutant

Acid trehalase was purified from the yeast suc2 deletion mutant. After hydrophobic interaction chromatography, the enzyme could be purified to a single band or peak by a further step of either polyacrylamide gel electrophoresis, gel filtration, or isoelectric focusing. An apparent molecular mass of 218,000 Da was calculated from gel filtration. Polyacrylamide gel electrophoresis of the purified enzyme in the presence of sodium dodecyl sulfate suggested a molecular mass of 216,000 Da. Endoglycosidase H digestion of the purified enzyme resulted after sodium dodecyl sulfate gel electrophoresis in one distinct band at 41,000 Da, representing the mannose-free protein moiety of acid trehalase. The carbohydrate content of the enzyme was 86%. Amino acid analysis indicated 354 residues/molecule of enzyme including 9 cysteine moieties and only 1 methionine. The isoelectric point of the enzyme was estimated by gel electrofocusing to be approximately 4.7. The catalytic activity showed a maximum at pH 4.5. The activity of the enzyme was not inhibited by 10 mM each of HgCl2, EDTA, iodoacetic acid, phenanthrolinium chloride or phenylmethylsulfonyl fluoride. There was no activation by divalent metal ions. The acid trehalase exhibited an apparent Km for trehalose of 4.7 +/- 0.1 mM and a Vmax of 99 mumol of trehalose min-1 X mg-1 at 37 degrees C and pH 4.5. The acid trehalase is located in the vacuoles. The rabbit antiserum raised against acid trehalase exhibited strong cross-reaction with purified invertase. These cross-reactions were removed by affinity chromatography using invertase coupled to CNBr-activated Sepharose 4B. Precipitation of acid trehalase activity was observed with the purified antiserum.     

Purification and characterization of heparinase from Flavobacterium heparinum

Heparinase (EC 4.2.2.7) isolated from Flavobacterium heparinum was purified to homogeneity by a combination of hydroxylapatite chromatography, repeated gel filtration chromatography, and chromatofocusing. Homogeneity was established by the presence of a single band on both sodium dodecyl sulfate and acid-urea gel electrophoretic systems. Amino acid analysis shows that the enzyme contains relatively high amounts of lysine residues (9%) consistent with its cationic nature (pI 8.5) but contains only 4 cysteine residues/polypeptide. The molecular weight of heparinase was estimated to be 42,900 +/- 1,000 daltons by gel filtration and 42,700 +/- 1,200 daltons by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme is very specific, acting only on heparin and heparan monosulfate out of 12 similar polysaccharide substrates tested. It has an activity maximum at pH 6.5 and 0.1 M NaCl and a stability maximum at pH 7.0 and 0.15 M NaCl. The Arrhenius activation energy was found to be 6.3 kcal/mol. However, the enzyme is very sensitive to thermal denaturation and loses activity very rapidly at temperatures over 40 degrees C. Kinetic studies of the heparinase reaction at 37 degrees C gave a Km of 8.04 X 10(-6) M and a Vm of 9.85 X 10(-5) M/min at a protein concentration of 0.5 microgram/ml. By adapting batch procedures of hydroxylapatite and QAE (quaternary aminoethyl)-Sephadex chromatography, gram quantities of heparinase that is nearly free of catalytic enzyme contaminants can be purified in 4-5 h.     

Purification and characterization of phosphatidylinositol-specific phospholipase C from bovine platelets

Phosphatidylinositol-specific phospholipase C was purified to homogeneity from soluble fraction of bovine platelets by ammonium sulfate fractionation, hydrophobic chromatography, DEAE ion exchange chromatography and gel filtration. The purified enzyme has a narrow pH optimum ranging from 6.5 to 7.5 and the molecular weight of the enzyme was estimated to be 143,000 by sodium dodecyl sulfate slab gel electrophoresis. The purified enzyme requires Ca2+ strictly for activity, which was markedly enhanced in the presence of arachidonate. No enhancement of the activity was observed in the presence of purified calmodulin. The activity was markedly inhibited in the presence of quinacrine but no inhibition by indomethacin was observed.     

Purification and some properties of skeletal muscle sarcolemma Ca2+-ATPase]

Ca2+-ATPase of skeletal muscle sarcolemma has been isolated and purified. It is prepared from salt extract of sarcolemma by ammonium sulfate fractionation and further purified by gel chromatography on Sepharose 4B. The purity of preparations was evaluated by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. It has been shown that Ca2+-ATPase possesses the same mobility as skeletal muscle myosin under gel chromatography on Sepharose 4B and the same mobility as myosin heavy chains in sodium dodecyl sulfate--polyacrylamide gel electrophoresis. Membrane protein binds to rabbit skeletal muscle actin, and this complex dissociates by ATP. Interaction with actin does not change Ca2+- or Mg2+-stimulated ATPase activity. Enzyme has only one pH optimum at 7,0-7,6. Membrane protein is highly specified to calcium--ATPase activity in the presence of Mn2+ is 10% and in the presence of Sr2+, Mg2+ or Co2+ are 3-5% of the activity in the presence of Ca2+. Other nucleoside triphosphate (UTP and ITP) are hydrolyzed at lower rates than is ATP.     

Purification and characterization of murine protoporphyrinogen oxidase

The penultimate enzyme of the heme biosynthetic pathway, protoporphyrinogen oxidase (EC 1.3.3.4), has been purified to apparent homogeneity from mouse liver mitochondria. The purification involves solubilization from mitochondrial membranes with sodium cholate followed by ammonium sulfate fractionation and gel filtration on a Sepharose CL-6B column. The eluate is adjusted to 0.67 M (NH4)2SO4 and loaded onto a phenyl-Sepharose column. After salt washes, the enzyme is eluted with 0.5% sodium cholate and 0.5% Brij 35. The final step is high-pressure ion-exchange chromatography on a DEAE-5PW column. The purified protein has a molecular weight of approximately 65,000 by gel filtration chromatography on Sepharose CL-6B in the presence of 0.5% sodium cholate. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis shows a single band corresponding to a molecular weight of 65,000. The absorption spectrum of the purified enzyme shows no evidence of a chromophoric cofactor. Purified protoporphyrinogen oxidase has a Km for protoporphyrinogen IX of 5.6 microM with a Vmax of 2300 nmol mg-1 h-1. It utilizes meso- and hematoporphyrinogen at about 10% the level of protoporphyrinogen. The pH optimum is broad with a maximum at 7.1. There is no stimulation or inhibition by any tested divalent cations, and sulfhydryl reagents have no inhibitory effect on the purified enzyme.     

Characterization of fructose 1,6-bisphosphatase from bakers' yeast

Active nonphosphorylated fructose bisphosphatase (EC 3.1.3.11) was purified from bakers' yeast. After chromatography on phosphocellulose, the enzyme appeared as a homogeneous protein as deduced from polyacrylamide gel electrophoresis, gel filtration, and isoelectric focusing. A Stokes radius of 44.5 A and molecular weight of 116,000 was calculated from gel filtration. Polyacrylamide gel electrophoresis of the purified enzyme in the presence of sodium dodecyl sulfate resulted in three protein bands of Mr = 57,000, 40,000, and 31,000. Only one band of Mr = 57,000 was observed, when the single band of the enzyme obtained after polyacrylamide gel electrophoresis in the absence of sodium dodecyl sulfate was eluted and then resubmitted to electrophoresis in the presence of sodium dodecyl sulfate. Amino acid analysis indicated 1030 residues/mol of enzyme including 12 cysteine moieties. The isoelectric point of the enzyme was estimated by gel electrofocusing to be around pH 5.5. The catalytic activity showed a maximum at pH 8.0; the specific activity at the standard pH of 7.0 was 46 units/mg of protein. Fructose 1,6-bisphosphatase b, the less active phosphorylated form of the enzyme, was purified from glucose inactivated yeast. This enzyme exhibited maximal activity at pH greater than or equal to 9.5; the specific activity measured at pH 7.0 was 25 units/mg of protein. The activity ratio, with 10 mM Mg2+ relative to 2 mM Mn2+, was 4.3 and 1.8 for fructose 1,6-bisphosphatase a and fructose 1,6-bisphosphatase b, respectively. Activity of fructose 1,6-bisphosphatase a was 50% inhibited by 0.2 microM fructose 2,6-bisphosphate or 50 microM AMP. Inhibition by fructose 2,6-bisphosphate as well as by AMP decreased with a more alkaline pH in a range between pH 6.5 and 9.0. The inhibition exerted by combinations of the two metabolites at pH 7.0 was synergistic.     

Purification, characterization and subcellular localization of pig liver alpha-L-iduronidase

alpha-L-Iduronidase was purified about 100,000-fold from pig liver by employing column chromatography on cellulose phosphate (P11), concanavalin A-Sepharose 4B, heparin-Sepharose 4B, Toyopearl HW-55, Sephadex G-100 and chelating Sepharose 6B charged with cupric ions. The molecular mass of the purified enzyme was estimated to be 70 kDa by Sephadex G-100 column chromatography. The purified enzyme gave a single band on disc polyacrylamide gel electrophoresis without using sodium dodecyl sulfate. However, two separate components of 70 kDa and 62 kDa appeared when it was analyzed by SDS/polyacrylamide gel electrophoresis. These 70-kDa and 62-kDa components were confirmed as alpha-L-iduronidase immunochemically. The isoelectric points of these enzymes were both 9.1 as measured by isoelectric focusing in a polyacrylamide gel containing ampholine and sucrose. The optimal pH and Km values were 3.0-3.5 and 65 microM 4-methylumbelliferyl-alpha-L-iduronide, respectively. The purified enzyme was stable in the pH range 3.5-6.0 under conditions with or without 0.5 M NaCl. However, in the presence of 0.5 M NaCl, it was unstable at pH 3.0. Moreover, it was conversely stabilized at pH 7.0 in the presence of 0.5 M NaCl. Immunohistochemically, the enzyme was found in the Kupffer cells and was abundant on their lysosomal membranes. In liver cells, however, the immunohistochemical reaction was weak.     

Human placenta alpha-N-acetylglucosaminidase: purification, characterization and demonstration of multiple recognition forms.

alpha-N-Acetylglucosaminidase (EC 3.2.1.50) was purified from human placenta by a four-step procedure including ammonium sulfate precipitation, affinity chromatography with immobilized antibodies against urinary alpha-N-acetylglucosaminidase, gel chromatography and discontinuous gel electrophoresis with a 30% recovery and 26 300-fold purification. Immunological methods revealed the contamination with about 10% non-alpha-N-acetylglucosaminidase protein. Isoelectric focusing led to a distribution of activity between 4.3 and 6.5 with maxima at pH 5.1 and pH 5.7. After treatment with neuraminidase, alpha-N-acetylglucosaminidase activity assembled at pH 5.7. The multiple isoelectric forms were endocytosed with different rates by cultured human skin fibroblasts. Placenta alpha-N-acetylglucosaminidase has an apparent molecular weight of 304 000 and contains 23.4% carbohydrate consisting of glucose, galactose, mannose, hexosamines and neuraminic acid. Gel electrophoresis in the presence of 0.1% sodium dodecylsulfate separated placenta alpha-N-acetylglucosaminidase into subunits with molecular weights of 86 500 and 81 000. The activity towards various substrates, the kinetics of hydrolysis, the pH optimum and the stability of the catalytic activity were assayed.     

Purification of a peptidylglycine alpha-amidating enzyme from transplantable rat medullary thyroid carcinomas

A peptidyl glycine alpha-amidating activity has been isolated from total tissue extracts of rat medullary thyroid carcinoma (MTC). Purification of the activity by ammonium sulfate fractionation, Sephacryl S-300 chromatography, and strong anion-exchange chromatography at pH 6.0 has resolved at least four peaks of activity. The activity associated with peak III has been further purified to apparent homogeneity by strong anion-exchange chromatography at pH 8.0. The purified peak III enzyme has an apparent molecular mass of 75,000 Da as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The identity of the 75,000-Da band as the alpha-amidating enzyme has been confirmed by recovery of activity from a nondenaturing polyacrylamide gel. The enzyme is catalytically active as a monomer, exhibits a pH optimum between 5.0 and 5.5, and has a turnover number of 300 min-1 for N-dansyl-Tyr-Val-Gly amidation at pH 5.5. The larger size, more acidic pH optimum, and higher specific activity distinguish the purified peak III rat MTC enzyme from the enzymes isolated from bovine and porcine pituitary or from frog skin.     

Purification of phospholipase D from citrus callus tissue

Phospholipase D in extracts of soluble proteins from callus cultures derived from cotyledons of Citrus sinensis (L.) Osbeck is activated by Ca2+ and anionic detergents and has a pH optimum of 6.5. The enzyme was purified 703-fold over the crude protein extract with a yield of 15% by ammonium sulfate precipitation, ion exchange chromatography, gel filtration, hydrophobic interaction chromatography, and preparative acrylamide gel electrophoresis. Preparative electrophoresis was carried out using conventional slab gel equipment and electroelution of the sliced gel. Analytical sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified phospholipase revealed two bands of the same staining intensity running at 94.2K and 90.5K.     

Purification of nitric oxide synthase from rat macrophages.

Nitric oxide (NO) synthase (EC 1.14.23) has been purified to apparent homogeneity from rat macrophages. The purification procedure involves affinity chromatography with adenosine 2',5'-diphosphate-agarose and gel filtration chromatography on a Superose 12 HR 10/30 column. The apparent molecular weight is 300,000 by gel filtration. On polyacrylamide gel electrophoresis in sodium dodecyl sulfate, the enzyme migrates as a single protein band with Mr = 150,000. The purified enzyme is colorless, and an absorption maximum is observed at 280 nm. The half-life of the enzyme activity is 6 h at pH 7.4 and 4 degrees C. The enzyme activity required the presence of NADPH, (6R)-5,6,7,8-tetrahydro-L-biopterin, and dithiothreitol. Although the cerebellar and endothelial enzyme require Ca2+ and calmodulin, these are not required by the macrophage enzyme. The macrophage nitric oxide synthase (an inducible enzyme) seems to be different from the cerebellar and endothelial enzyme (a constitutive enzyme).     

Purification and characterization of glycerol-3-phosphate dehydrogenase (flavin-linked) from rat liver mitochondria

We have purified the membrane-intrinsic glycerol-3-phosphate dehydrogenase from both normal and hyperthyroid rat liver mitochondria by extraction with Triton X-100, hydrophobic affinity chromatography, ion exchange chromatography, gel filtration, and FAD-linked Sepharose 4B affinity chromatography. The yields in both cases were over 20%, and purification ranged from 800- to 650-fold in mitochondria from hyperthyroid and normal rats, respectively. The final preparations appeared to be greater than 95% pure by polyacrylamide gel electrophoresis in the presence or absence of sodium dodecyl sulfate. The pure enzyme focused at pH 5.5 and produced a biphasic thermal inactivation plot at 50 degrees C. The holoenzyme was found to have a molecular mass of 250,000 daltons on gel filtration. The subunit molecular mass was found to be 74,000 daltons +/- 3,000 by sodium dodecyl sulfate-gel electrophoresis and high-performance liquid chromatography gel filtration in 0.1% sodium dodecyl sulfate. 1 mol of the holoenzyme preparation contains 1.1 mol of non-heme iron and 0.7-0.9 mol of noncovalently bound FAD. The absorption spectrum has a maximum at 375 nm and a shoulder at 450 nm which is bleached on treatment with sodium dithionite. The enzymatic reaction is competitively inhibited by glyceraldehyde 3-phosphate, dihydroxyacetone phosphate, phosphoenolpyruvate, and phosphoglycolic acid. The apparent Km for DL-alpha-glycerol 3-phosphate and noncovalently bound FAD were found to be 6 mM and 7 microM, respectively.     

Purification and properties of a nad: 3α-hydroxy-5α-pregnan-20-one-oxidoreductase from rat liver microsomes

From rat liver microsorties a NAD: 3α-hydroxy-5α-pregnan-20-one oxidoreductase was isolated and purified up to a specific activity of 73 nmol/min.mg by affinity chromatography and DEAE-cellulose chromatography. Various Km-values have been determined. The enzyme exhibits highest affinity for 5α-pregnane-3,20-dione and NADH. The 3-oxo group of 5α-dihydrocortisone (17, 21-dihydroxy-5α-pregnane-3,11,20-trione) was not reduced by the purified enzyme preparation and NADH and no dehydrogenation with NAD was observed of 3α, 11β, 17, 21-tetrahydroxy-5α-pregnan-20-one. The optimal pH for the hydrogenation of the 3-oxo group was at pH 5.3 and for the dehydrogenation at pH 8.9. Disc gel electrophoresis in presence of 0.1% sodium dodecylsulfate yielded a homogeneous preparation.     

Prenyltransferase from Saccharomyces cerevisiae. Purification to homogeneity and molecular properties.

Prenyltransferase (EC 2.5.1.1) has been purified to homogeneity from the supernatant fraction of yeast by ammonium sulfate fractionation, diethylaminoethyl-cellulose and hydroxylapatite chromatography, and column isoelectric focusing techniques. The active enzyme from isoelectric focusing columns emerged as a single symmetrical peak with specific activities 15- to 35-fold higher than previously reported preparations. The enzyme was found to be homogeneous by continuous polyacrylamide gel electrophoresis at pH 8.4 and discontinuous polyacrylamide gel electrophoresis at pH 6.9 as well as sodium dodecyl sulfate polyacrylamide electrophoresis at pH 7.0. By means of gel chromatography and sodium dodecyl sulfate polyacrylamide gel electrophoresis, the protein was shown to be a dimer with a molecular weight of 84,000 plus or minus 10%. The isoelectric point of the enzyme was determined to be 5.3. The enzyme synthesizes farnesyl and geranylgeranyl pyrophosphates from dimethylallyl, geranyl, and farnesyl pyrophosphates. Michaelis constants for the enzyme were 4, 8, and 14 mu M for isopentenyl, dimethylallyl, and geranyl pyrophosphates, respectively.     

Biochemical and immunological studies of purified mouse beta-galactosidase.

Beta-Galactosidase (EC 3.2.1.23) has been purified from the livers of C57BL/6J mice. The enzyme migrated as a single band of protein on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. The molecular weight of the denatured and reduced enzyme was 63,000. The native form of beta-galactosidase appeared to be a tetramer of 240,000 at pH 5.0, which was reversibly dissociated at alkaline pH to a dimer with apparent molecular weight of 113,000. Multiple charge isomers of beta-galactosidase were resolved by polyacrylamide gel electrophoresis and ion exchange chromatography. Treatment of beta-galactosidase with neuraminidase markedly reduced its electrophoretic mobility. Purified enzyme as well as crude liver extract hydrolyzed p-nitrophenyl-beta-D-fucoside at one-tenth the rate of hydrolysis of the beta-galactoside. Antiserum to the purified enzyme precipitated the major portion of beta-galactosidase activity of mouse liver, brain, and kidney. This antiserum cross-reacts with beta-galactosidases from rat and Chinese hamster, but not with human, porcine, or bovine beta-galactosidase.     

Purification and partial characterization of cysteine-glutamate transaminase from rat liver.

Cysteine-glutamate transaminase (cysteine aminotransferase; EC 2.6.1.3) has been purified 149-fold to an apparent homogeneity giving a specific activity of 2.09 IU per milligram of protein with an overall yield of 15%. The isolation procedures involve the preliminary separation of a crude rat liver homogenate which was submitted sequentially to ammonium sulfate fractionation, TEAE-cellulose column chromatography, ultrafiltration, and isoelectrofocusing. The final product was homogenous when examined by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS). A minimal molecular weight of 83 500 was determined by Sephadex gel chromatography. The molecular weight as estimated by polyacrylamide gel electrophoresis in the presence of SDS was 84 000. The purified enzyme exhibited a pH optimum at 8.2 with cysteine and alpha-ketoglutarate as substrates. The enzyme is inactivated slowly when kept frozen and is completely inactivated if left at room temperature for 1 h. The enzyme does not catalyze the transamination of alpha-methyl-DL-cysteine, which, when present to a final concentration of 10 mM, exhibits a 23.2% inhibition of transamination of 30 mM of cysteine. The mechanism apparently resembles that of aspartate-glutamate transaminase (EC 2.6.1.1) in which the presence of a labile hydrogen on the alpha-carbon in the substrate is one of the strict requirements.     

Isolation of a new ribonuclease from fruiting bodies of the silver plate mushroom Clitocybe maxima

A ribonuclease, with an N-terminal sequence exhibiting some homology to ribonuclease from Pleurotus ostreatus (Family Pleurotaceae), has been purified from fruiting bodies of the silver plate mushroom Clitocybe maxima (Family Tricholomataceae). However, there is little resemblance between the N-terminal sequences of ribonucleases from various Pleurotus species, and a lesser extent of resemblance between ribonucleases from C. maxima and Pleurotus tuber-regium. No structural relationship exists between ribonuclease from C. maxima, and those from Volvariella volvacea, Lentinus edodes and Irpex lacteus. The purification protocol involved ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, ion exchange chromatography on CM-Sepharose, and fast protein liquid chromatography on Superdex 75. The ribonuclease was unadsorbed on DEAE-cellulose and adsorbed on Affi-gel blue gel and CM-Sepharose. It exhibited a molecular mass of 17.5 kDa in both gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It manifested roughly the same ribonucleolytic potency toward poly A and poly G followed by poly U. Its activity toward poly C was, by comparison, meager. The temperature and pH required for its optimal activity were, respectively, 70 degrees C and 6.5-7.0.