Purine nucleoside phosphorylases purified from rat liver and Novikoff hepatoma cells.

Purine nucleoside phosphorylase (PNP) was purified from rat hepatoma cells and normal liver tissue utilizing the techniques of ammonium sulfate fractionation, heat treatment, ion-exchange and molecular exclusion chromatography, and polyacrylamide gel electrophoresis. Homogeneity was established by disc gel electrophoresis in the presence and absence of sodium dodecyl sulfate. Purified rat hepatoma and liver PNPs appeared to be identical with respect to subunit and native molecular weight, substrate specificity, heat stability, kinetics and antigenic identity. A native molecular weight of 84,000 was determined by gel filtration. A subunit molecular weight of 29,000 was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A single isoelectric point was observed at pH 5.8, and the pH optimum was 7.5. Inosine, guanosine, xanthosine, and 6-mercaptopurine riboside were substrates for the enzymes. The apparent K_m for both inosine and guanosine was about 1.0 × 10^?4m and for phosphate was 4.2 × 10^?4m. Hepatoma and liver PNP showed complete cross-reactivity using antiserum prepared against the liver enzyme.     

Rabbit brain purine nucleoside phosphorylase. Physical and chemical properties. Inhibition studies with aminopterin, folic acid and structurally related compounds.

Rabbit brain purine nucleoside phosphorylase used in this study was purified 6000-fold to apparent homogeneity and a specific activity or 50 μmol min^?1 mg ^?1 protein. A molecular weight of 70.000 daltons was determined for the native enzyme by gel filtration on Sephadex. Electrophoresis on polyacrylamide gel, in presence of sodium dodecyl sulfate, gave a subunit molecular weight of 34,500 daltons, suggesting that the enzyme is dimeric with, probably, identical subunits. The relationship of the structure of certain biologically active substances to their inhibitory action on the enzyme was examined. Folic acid and the compound d,l-6-methyl 5,6,7,8-tetrahydropterine, with similar substituents on their primary ring structure, were competitive inhibitors of the enzyme. The inhibition constants calculated were 3.37 × 10^?5M for folic acid and 3.80 × 10^?5m for d,l-6-methyl 5,6,7,8-tetrahydropterine. Aminopterin and the purine analog 8-aza-2,6-diaminopurine, with similar substituents on their primary ring structure, were noncompetitive inhibitors of the enzyme. Their respective inhibition constants were 1.50 × 10^?4 and 1.95 × 10^?4m. Erythro-9-(2-hydroxy-3-nonyl) adenine, an adenosine deaminase inhibitor, was also examined for inhibitory potency with mammalian purine nucleoside phosphorylase, and was observed to be a competitive inhibitor of this enzyme, with an inhibition constant of 1.90 × 10^?4m. The Michaelis constant for the substrate guanosine was near 6.0 × 10^?5m. Physical probe of the nature of the functional groups which participate in enzymic catalysis implicated both histidine and cysteine as the essential catalytic species. Photooxidation studies suggested a pH-dependent sensitivity of an essential catalytic group, and its probable location at the active site.     

Studies on NADPH-cytochrome c reductase I: Isolation and several properties of the crystalline enzyme from ale yeast.

NADPH-cytochrome c reductase has been isolated from a top-fermenting ale yeast, Saccharomyces cerevisiae (Narragansett strain), after ca. a 240-fold purification over the initial extract of an acetone powder, with a final specific activity (at pH 7.6, 30 °C) of ca. 150 μmol cytochrome c reduced min^?1mg^?1 protein. The preparation appears to be homogeneous by the criteria of: sedimentation velocity; electrophoresis on cellulose acetate in buffers above neutrality; and by polyacrylamide gel electrophoresis. Although the reductase appeared to partially separate into species “A” and “B” on DEAE-cellulose at pH 8.8, the two species have proven to be indistinguishable electrophoretically (above pH 8) and by sedimentation. By sedimentation equilibrium at 20 °C, a molecular weight of ca. 6.8 (± 0.4) × 10⁴ was obtained with use of a $\text{V}\text{?}{}_{\mathrm{20\; °}}\text{= 0.741}$ calculated from its amino acid composition. After disruption in 4 m guanidinium chloride- 10 mm dithioerythritol- 1 mm EDTA, pH 6.4 at 20 °C, an $\text{M}\text{?}{}_{\mathrm{<mtext>r</mtext>}}$ of 3.4 (± 0.1) × 10⁴ resulted, which points to a subunit structure of two polypeptide chains per mole. Confirmatory evidence of the two-subunit structure with similar, if not identical, polypeptide chains was obtained by polyacrylamide gel electrophoresis in dodecyl-sulfate, after disruption in 4 m urea and 2% sodium dodecyl sulfate, and yielded a subunit molecular weight of ca. 4 × 10⁴. Sulfhydryl group titration with 4,4′-dithiodipyridine under acidic conditions revealed one sulfhydryl group per monomer, which apparently is necessary for the catalytic reduction of cytochrome c. NADPH, as well as FAD, protects this-SH group from reaction with 5,5′-dithiobis (2-nitrobenzoate). The visible absorption spectrum of the oxidized enzyme (as prepared) has absorption maxima at 383 and 455 nm, typical of a flavoprotein. Flavin analysis (after dissociation by thermal denaturation of the “A” protein) conducted fluorometrically, revealed the presence of 2.0 mol of FAD per 70,000 g, in confirmation of the deduced subunit structure. The identity of the FAD dissociated from either “A” or “B” protein was confirmed by recombination with apo-d-amino acid oxidase and by thin-layer chromatography. A kinetic approach was used to estimate the dissociation constant for either FAD or FMN (which also yields a catalytically active enzyme) to the apoprotein reductase at 30 °C and pH 7.6 (0.05 m phosphate) and yielded values of 4.7 × 10^?8m for FAD and 4.4 × 10^?8m for FMN.     

Studies on (K + H)-ATPase. V. Chemical composition and molecular weight of the catalytic subunit

(1) A (K⁺ + H⁺)-ATPase preparation from porcine gastric mucosa is solubilized in sodium dodecyl sulfate, and is subjected to gel filtration. (2) A main subunit fraction is obtained, which is a protein carbohydrate lipid complex, containing 88% protein, 7% carbohydrate and 5% phospholipid. The detailed composition of the protein and carbohydrate moieties are reported. (3) Sedimentation analysis of the subunit preparation, after detergent removal, reveals no heterogeneity, but the subunits readily undergo aggregation. (4) Acylation of the subunit preparation with citraconic anhydride causes a clear shift of the band obtained after SDS gel electrophoresis, but the absence of broadening and splitting of the band pleads against subunit heterogeneity. (5) Treatment of the subunit preparation with dansyl chloride indicates that the NH₂ terminus is blocked, which favors the assumption of homogeneity of the protein. (6) Binding studies with concanavalin A indicate that at least 86% of the subunit preparation is composed of glycoprotein. (7) These findings, taken together, strongly suggest that there is a single subunit which is a glycoprotein and which represents the catalytic subunit of the enzyme. From sedimentation equilibrium analysis a molecular mass value of 119 kDa (S.E. 3, n = 6) is calculated for protein + carbohydrate and of 110 kDa (S.E. 3, n = 6) for protein only. (8) In combination with the molecular mass of 444 kDa (S.E. 10, n = 4) obtained for the intact enzyme by radiation inactivation we conclude that the enzyme appears to be composed of a homo-tetramer of catalytic subunits.     

Some properties of a glycoprotein with calcium binding ability found in human biological fluids in macroglobulinaemia IgM

An acidic glycoprotein with calcium-binding properties was isolated from the urine of patients with severe macroglobulinaemia IgM. The molecular weight of this protein determined by Sephadex gel filtration was found to be 62 000 ± 2800 in Tris · HCl buffer and 21 000 ± 1 000 in 6 M guanidine · HCl. The amino acid and carbohydrate composition of the isolated glycoprotein is presented. Electrophoretic migration of this protein was observed to be greatly affected by calcium ions present in the buffer in a concentration of 10^?3 M. At least two sets of binding sites seem to participate in binding calcium. The values 2.2 · 10⁶ M^?1 for the apparent association constant and 4.4 · 10^?4 mol of Ca²⁺ bound per g of protein for high affinity binding sites were estimated, on the basis of data from the equilibrium dialysis. The origin possible biological role of this protein is discussed.     

An aminopeptidase from Agave americana,isolation and physical characterization

A new aminopeptidase was isolated from Agave americana by fractionation, chromatography and gel filtration. The mw of the enzyme, determined by different procedures was 86000 ± 1500; the enzyme had a sedimentation coefficient of 4.96 S, a diffusion coefficient of 5.2 × 10^?7 cm²/sec, a Stokes radius of 3.8 nm, a partial specific volume of 0.733 cm³/g, a frictional ratio of 1.40, a molecular absorbancy index at 280 nm of 8.36 × 10⁴, an isoelectric point of 4.53 and contained 1.25% carbohydrate. The amino acid composition of the enzyme was determined and the aminoterminal residue was identified as lysine whilst the carboxylterminal residue was either leucine or isoleucine. No subunit structure was observed for the enzyme.     

Flotation equilibrium of serum low density lipoproteins

The molecular weight of human serum low density lipoprotein (LDL) was determined for the first time by sedimentation equilibrium or, more accurately, flotation equilibrium, in high concentration of KBrNaBr containing Tris-Cl buffer plus EDTA (density = 1.20 – 1.49 g/ml). Assuming both the molecular weight and the partial specific volume were unknown, the results at different densities gave a value of 2.87 ± 0.12 × 10⁶ for the molecular weight and 0.965 ± 0.014 ml/g for the partial specific volume.     

Purification and partial characterization of alcohol dehydrogenase from wheat.

Further support for hypotheses proposed earlier for the genetic control and subunit composition of the alcohol dehydrogenase of Triticum has been obtained through the purification and partial characterization of the enzyme. The alcohol dehydrogenase of the wheat T. monococcum was purified 103-fold to a specific activity of 55,900 units/mg. Purification was achieved using streptomycin sulfate precipitation, gel filtration chromatography, DEAE-cellulose anion-exchange chromatography, and preparative isoelectric focusing. The native enzyme has a molecular weight of 116,000 and a dimeric subunit structure. The apparent Michaelis constants are 1.2 × 10^?2m for ethanol and 1 × 10^?4m for NAD. The substrate specificity of wheat alcohol dehydrogenase differs significantly from the substrate specificities of the enzymes of horse and yeast.     

Purification and Some Properties of Saccharomyces logos α-Glucosidase

α-Glucosidase was purified from Saccharomyces logos by precipitation with ethanol, and chromatographies on Sephadex G–200, DEAE-Sephadex, DEAE-ceiluiose and Duolite A–2. The purified α-glucosidase was homogeneous on ultracentrifugation and zone electrophoresis using cellulose acetate membrane. The sedimentation coefficient was calculated to be 9.6 S. The molecular weight was estimated to be approximately 2.7 × 10⁵ by gel-filtration technique.

The optimum pH was found to be in the range of 4.6~5.0, and the optimum temperature was 40°C. The enzyme exhibited higher hydrolytic activity toward maltose rather than toward phenyl-α-glucoside and turanose, and no activity toward sucrose.

The enzyme was a glycoprotein containing carbohydrate of about 50%.     

The atypical RNA components of cytoplasmic ribosomes from Crithidia fasciculata

RNA extracted by cold phenol from the large cytoplasmic ribosomal subunit of the trypanosomatid flagellate Crithidia fasciculata and analyzed by polyacrylamide gel electrophoresis at 4 °C consisted of one species with a molecular weight of 1.3 × 10⁶ (relative to ribosomal RNA from E. coli MRE 600). When extracted with hot phenol (65 °C), the large ribosomal subunit gave rise to two components with molecular weights of 0.72 and 0.56 × 10⁶. On heating for 60 s, followed by rapid cooling, the single cold-phenol-extracted 1.30 × 10⁶-dalton species completely dissociated into two components of molecular weights 0.72 and 0.56 × 10⁶, present in equimolar amounts. When analyzed by polyacrylamide-agarose gel electrophoresis in the presence of SDS, RNA extracted by cold phenol from the large cytoplasmic ribosomal subunit consisted of three components of molecular weights 1.3, 0.72, and 0.56 × 10⁶, present in apparently equimolar amounts. RNA from the small cytoplasmic ribosomal subunit consisted of one species with a molecular weight of 0.84 × 10⁶, independent of extraction or analytical conditions. It is proposed that under high salt and low temperature conditions, the large ribosomal RNA molecule is held together by its secondary structure, and that denaturing extraction or analytical conditions reveal an otherwise “hidden” lesion present in the molecule in vivo.     

Molecular interactions between ribosomal proteins: a study of the S6-S18 interaction.

The proteins S6 and S18 from the 30 S ribosomal subunit of Escherichia coli were isolated to a purity of greater than 95%, characterized in solution, and investigated by sedimentation equilibrium for possible intermolecular interactions in a dilute salt reconstitution buffer. It was observed that neither protein S6 nor S18 has a tendency to self-associate in the concentration range studied. An analysis of solution mixtures containing proteins S6 and S18 revealed a species of molecular weight greater than either of the proteins. Proteins S6 and S18 were found to interact with an equilibrium constant of association of 6.6 ± 4.2 × 10⁴m^?1 at 3 °C with a Gibbs free energy of interaction, ΔG ° = ?6.1 kcal/mol. These data are part of those collected to help in building a map of the energetics in the 30 S ribosomal subunit, which provides for the stabilization of the structure.     

Quaternary structure of the isolated restriction endonuclease EndoR.Bgl I from Bacillus globigii as revealed by electron microscopy.

The restriction endonuclease EndoR · BglI was purified nearly to homogeneity. BglI samples, when negatively stained with 4% uranyl acetate, show two different particle projections in the electron microscope. Projection A has an outer diameter of 22.5 ± 0.8 nm and is composed of six intensity maxima arranged in a ring; the centre of the ring exhibits slightly visible additional substructures. Projection B is also a ring; its outer diameter is 23.8 ± 0.7 nm; it does not show detailed fine structure, aside from a probable 10-fold rotational symmetry. Variations of the negative staining technique (single carbon layer, 2% uranyl acetate; ‘sandwich’ preparation with 4% uranyl acetate) revealed additional fine structural details for both projections. From the electron microscopic observations, a model of the enzyme particle was developed containing 20 identical, biologically active monomers of molecular weight around 61,000, arranged as a pentagonal dodecahedron. Tilting experiments established this structure decisively by interconversion of the different appearances of given particles in the expected way. By sodium dodecyl sulphate/polyacrylamide gel electrophoresis, polyacrylamide gel electrophoresis in a continuous molecular sieve gradient and evaluation of negatively stained enzyme particles, a molecular weight of the monomer of 61,000 was estimated, resulting in a total enzyme particle molecular weight of 1.2 × 10⁶ also determined by linear sucrose density-gradient centrifugation.     

Types of Amylases in Rice Grains

The subunit structure of phenoloxidase from the larvae of housefly, Musca domestica vicina Maquart, was investigated by urea treatment, SDS-polyacrylamide gel electrophoresis and electron microscopy. Phenoloxidase was dissociated into a single type of subunit (MW 6.5 × 10⁴) by 5 m urea treatment. A similar subunit (MW 6.1 × 10⁴) was also detected by SDS-polyacrylamide gel electrophoresis. Furthermore, four additional protein bands with molecular weights of 4.8 × 10⁴, 3.5 × 10⁴, 2.4 × 10⁴ and 1.2 × 10⁴ were detected by SDS-polyacrylamide gel electrophoresis by extensive incubation of phenoloxidase with SDS. A cylindrical molecular structure was deduced from the electron microscopic analysis. The outside and inside diameter and the height of phenoloxidase molecule were evaluated to be 100, 50 and 70 Å, respectively.     

Molecular weight and symmetry of the pyruvate dehydrogenase multienzyme complex of Escherichia coli.

The molecular weight and polypeptide chain stoichiometry of the native pyruvate dehydrogenase multienzyme complex from Escherichia coli were determined by independent techniques. The translational diffusion coefficient (D^o_20,w) of the complex was measured by laser light intensity fluctuation spectroscopy and found to be 0.90 (±0.02) × 10^?11m²/s. When this was combined in the Svedberg equation with the measured sedimentation coefficient (s^o_20,w = 60.2 (±0.4) S) and partial specific volume ($\text{v}\text{?}\text{= 0.735 (±0.01)}\text{ml/g}$), the molecular weight of the intact native complex was calculated to be 6.1 (±0.3) × 10⁶. The polypeptide chain stoichiometry (pyruvate decarboxylase: lipoate acetyltransferase: lipoamide dehydrogenase) of the same sample of pyruvate dehydrogenase complex was measured by the radioamidination technique of Bates et al. (1975) and found to be 1.56:1.0:0.78.From this stoichiometry and the published polypeptide chain molecular weights estimated by sodium dodecyl sulphate/polyacrylamide gel electrophoresis, a minimum chemical molecular weight of 283,000 was calculated. This structure must therefore be repeated approximately 22 times to make up the native complex, a number which is in good agreement with the expected repeat of 24 times if the lipoate acetyltransferase core component has octahedral symmetry. It is consistent with what appears in the electron microscope to be trimer-clustering of the lipoate acetyltransferase chains at the corners of a cube. It rules out any structure based on 16 lipoate acetyltransferase chains comprising the enzyme core.The preparation of pyruvate dehydrogenase complex was polydisperse: in addition to the major component, two minor components with sedimentation coefficients (s^o_20,w) of 90.3 (±0.9) S and 19.8 (±0.3) S were observed. Together they comprised about 17% of the total protein in the enzyme sample. Both were in slowly reversible equilibrium with the major 60.2 S component but appeared to be enzymically active in the whole complex reaction. The faster-sedimenting species is probably a dimer of the complex, whereas the slower-sedimenting species has the properties of an incomplete aggregate of the component enzymes of the complex based on a trimer of the lipoate acetyltransferase chain.     

Ribulose 1,5-bisphosphate carboxylase from the halophilic cyanobacterium Aphanothece halophytica

Various structural and functional properties of ribulose 1,5-bisphosphate carboxylase/ oxygenase (RuBisCO) isolated from the halophilic cyanobacterium (blue-green alga) Aphanothece halophytica were reexamined. The ready dissociation of this algal RuBisCO during sedimentation in a linear sucrose density gradient was observed. Low NaCl concentrations promote the dissociation of small subunit (B) from the original native enzyme molecule as evidenced by the sucrose density gradient centrifugation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. It is thus possible that the intracellular osmoticum of A. halophytica might influence the structural integrity and activity of RuBisCO. The low residual carboxylase activity ascribed to the catalytic core, an oligomer form of the large subunit (A) apparently deficient in small subunit (B), was found to be markedly stimulated by a protein component which appears identical to subunit B. The purification and structural characterization of the catalytic core and subunit B were attempted by step-wise column chromatography on DEAE-cellulose, Utrogel AcA 34, Sephadex G-75, and hydroxylapatite, and at the final stage each component was purified to near homogeneity, although the catalytic core is still associated with a small quantity of subunit B. The addition of subunit B to the catalytic core does not alter the K_m (HCO₃^?, RuBP) values, but V_max values are markedly enhanced. Sucrose density gradient centrifugation gave a value of 16 S for the catalytic core. The molecular weights of the monomeric forms of the catalytic core (subunit A) and subunit B were 5.0 × 10⁴ and 1.4 × 10⁴, respectively.     

The flexibility of low molecular weight double-stranded dna as a function of length: II. Light scattering measurements and the estimation of persistence lengths from light scattering,sedimentation and viscosity

In the preceding paper are described the isolation and physical characterization of seven narrowly disperse fractions of calf thymus DNA in the molecular weight range 0.3 to 1.3 × 10⁶ daltons. Herein, we have determined by light scattering the molecular weights and root mean square radii of these fractions in a solvent comprising 0.2 M NaCl, 2 mM EDTA, 2m MNa-PO₄, pH 7. Measurements were made in a modified Wippler—Scheibling photometer to a 20° lower limit of scattering angle on solutions rendered virtually dust-free by procedures described. The optical aniso tropics of the DNA fractions were measured permitting the experimental molecular weights and root mean square radii to be corrected to their true values. From these values, with appropriate polydispersity corrections, we calculate a Kratky—Porod persistence length, a, of 54.0 ± 5.6 nm which is invariant over the molecular weight range examined. From the sedimentation coefficients (preceding paper) and the theory of Yamakawa and Fujii, we calculate a to be 66 nm, a value found to apply equally well to several DNA samples of various origins whose sedimentation rates are known in the molecular weight range from about 4 × 10⁴ to 10⁸ daltons. Similarly, from the intrinsic viscosities and the theory of Yamakawa and Fujii, we calculate a to be 59 nm, which again adequately applies to a number of DNA samples whose viscosities have been measured by other workers in the molecular weight range 3 × 10⁵ to 10⁸ daltons. The Flory—Mandelkern parameter, β, was found to vary with molecular weight in the manner predicted by the theory of Yamakawa and Fujii. The average value of a from the three sets of measurements is 60 ± 6 nm, which we believe applies to double-stranded DNA molecules, independent of chain length, over the whole range of molecular weights for which reliable data exist.     

Homology of plasmids in strains of unicellular Cyanobacteria.

Six strains of unicellular cyanobacteria were examined for the presence of plasmids. Analysis of lysates of these strains by CsCl-ethidium bromide density centrifugation yielded a major chromosomal DNA band and a minor band containing covalently closed circular plasmid DNA, as shown by electron microscopy and agarose gel electrophoresis. The sizes of the various plasmid species were determined; in each of the Synechococcus strains 6301, 6707, and 6908 two plasmid species were found with molecular weights of 5.3 × 10⁶ and 32.7 × 10⁶. Synechococcus strain 7425 had two plasmids of molecular weight 5.4 × 10⁶ and 24 × 10⁶. Synechococcus strain 6312 and Synechocystis strain 7005 each contained one plasmid species with molecular weight of 15.9 × 10⁶ and 2.0 × 10⁶, respectively. Restriction enzyme analysis revealed identical cleavage patterns for the plasmids of identical molecular weight.     

Water-Soluble Glycoconjugates of Vegetables I. Isolation and Properties of Arabinogalactan-Proteins from Cabbage

We present a new method for isolating and purifying water-soluble arabinogalactan-proteins from cabbage and give their chemical properties. The water-soluble nondialyzable material from fresh cabbage was separated into three fractions (A-I, II, and III) by gel filtration on Sepharose CL-4B. A-I and A-II can be purified by HPLC. Borate is necessary to avoid formation of insoluble aggregates during isolation and purification. The molecular weights of A-I, II, and III were estimated to be 4.0×10⁵, 1.0×10⁵, and 1.0~4.0×10⁴, respectively. A-I and A-II are arabinogalactan-proteins with different carbohydrate/protein ratios: 5.5/1 for A-I and 11.4/1 for A-II. The carbohydrate moieties of A-I and A-II were both arabino-3,6-galactans having d-galactose/l-arabinose ratios of 1.9/1 and 1.5/1, respectively. The amino acid composition indicates an abundance of hydroxyproline, serine, threonine and alanine, the sum of which amounted to about 50% of the total amino acids. A-I contained 1.5 times more hydroxyproline (20%) than A-II (14%), while A-II contained higher proportions of serine, glycine, and alanine. A-III was not a glycoprotein but was a mixture of carbohydrate and polypeptides.     

The free solution mobility of DNA

The free solution mobility of DNA has been measured by capillary electrophoresis in the two buffers most commonly used for DNA gel electrophoresis, Tris-borate-EDTA (TBE) and Tris-acetate-EDTA (TAE). The capillaries were coated with polymers of either of two novel acrylamide monomers, N-acryloylaminoethoxyethanol or N-acryloylaminopropanol, both of which are stable at basic pH and effectively eliminate the electroendosmotic mobility due to the capillary walls. The free solution mobility of DNA in TAE buffer was found to be (3.75 ± 0.04) × 10⁻⁴ cm² V⁻¹ s⁻¹ at 25°C, independent of DNA concentration, sample size, electric field strength, and capillary coating, and in good agreement with other values in the literature. The free solution mobility was independent of DNA molecular weight from ∼ 400 base pairs to 48.5 kilobase pairs, but decreased monotonically with decreasing molecular weight for smaller fragments. Surprisingly, the free solution mobility of DNA in TBE buffer was found to be (4.5 ± 0.1) × 10⁻⁴ cm² V⁻¹ s⁻¹, about 20% larger than observed in TAE buffer, presumably because of the formation of nonspecific borate-deoxyribose complexes. © 1997 John Wiley & Sons, Inc. Biopoly 42: 687–703, 1997     

Aggregation of tyrocidine in aqueous solutions.

The formation of aggregates of tyrocidine B at 4°C and 20°C in aqueous solutions was studied by means of light scattering and fluorescent techniques. The apparent weight molecular weight of tyrocidine B aggregates was found to be 36,000 at 4°C and 28,800 at 20°C. Fluorescence titration experiments with dansyl-chloride resulted in an aggregational number of 31 (4°) and 28 (20°) indicating that one molecule of dye is bound per monomer of molecular weight 1,200. From a Scatchard plot apparent association constants of 1.22 × 10⁵ M (4°) and 0.95 × 10⁵ M (20°) were calculated. From the angular dependence of scattered intensity the radii of gyration were determined to be 60 Å and 58 Å, respectively.