SULFHYDRYL AND DISULFIDE GROUPS OF PROTEINS : IV. SULFHYDRYL GROUPS OF THE PROTEINS OF MUSCLE

1. In the denatured proteins of skeletal muscle, the ratio of SH to S-S groups is higher than in the mixed denatured proteins of other tissues, with a single exception—the proteins of the crystalline lens. 2. The number of active SH groups in the proteins of minced muscle or in any of the protein fractions of muscle is only a fraction of the number found after the proteins have been treated with a denaturing agent. 3. The SH groups of the native proteins of muscle are activated by a rise in pH. 4. The relation between pH and number of active SH groups in the proteins of minced muscle and in the various protein fractions of muscle shows that little, if any, denatured protein is present in minced muscle.     

SULFHYDRYL AND DISULFIDE GROUPS OF PROTEINS : I. METHODS OF ESTIMATION

1. Methods have been described for reducing protein S-S groups, for oxidizing protein SH groups, and for estimating protein S-S and SH groups. 2. It has been found necessary in estimating the cystine content of proteins by the Folin-Marenzi method to take into account any cysteine that may be present. 3. A method for estimating the cysteine content of proteins has been described. 4. With these methods, estimations have been made of the S-S and SH groups and of the cystine and cysteine contents of a number of proteins. 5. In a denatured, but unhydrolyzed protein, the number of S-S and SH groups is equivalent to the quantity of cystine and cysteine found in the protein after hydrolysis.     

RELATION BETWEEN WATER PERMEABILITY AND INTEGRITY OF SULFHYDRYL GROUPS IN MALIGNANT AND NORMAL CELLS

When malignant cells, animal and human, were exposed in vitro to solutions of heavy metals or other selected compounds, three types of cell blebs were produced: (1) acentric blebs, arising from one side of the cell, e. g., by chlormerodrin, meralluride sodium, mercuric chloride; (2) symmetrical blebs; which completely enveloped the cell, e. g., by strong silver protein, auric chloride, p-chloromercuribenzoate; (3) scallop blebs, numerous small spherical elevations which completely covered the cell, e.g., by N-ethyl-maleimide, trivalent arsenicals, iodoacetamide. As indicated by vital stains and morphologic appearance, the blebs arose in healthy cells. They also can be made to appear in vivo in ascites tumor cells by intraperitoneal administration of a blebbing agent. All the bleb-producing chemicals have the property of reacting with protein-sulfhydryl groups by alkylation, oxidation or mercaptide formation. The three bleb types have been induced in 8 mouse and 2 rat ascites tumor cells; in 4 human and 1 mouse malignant cell lines; and in 3 normal cell lines grown in tissue culture. In contrast, cells from normal solid tissues of liver, lung, spleen, kidney, testis and brain from mouse, rat and rabbit failed to produce blebs. A possible interpretation for these observations is presented.     

STUDIES IN THE PHYSICAL CHEMISTRY OF THE PROTEINS : II. THE RELATION BETWEEN THE SOLUBILITY OF CASEIN AND ITS CAPACITY TO COMBINE WITH BASE. THE SOLUBILITY OF CASEIN IN SYSTEMS CONTAINING THE PROTEIN AND SODIUM HYDROXIDE.

1. The solubility in water of purified, uncombined casein has previously been reported to be 0.11 gm. in 1 liter at 25°C. This solubility represents the sum of the concentrations of the casein molecule and of the soluble ions into which it dissociates. 2. The solubility of casein has now been studied in systems containing the protein and varying amounts of sodium hydroxide. It was found that casein forms a well defined soluble disodium compound, and that solubility was completely determined by (a) the solubility of the casein molecule, and (b) the concentration of the disodium casein compound. 3. In our experiments each mol of sodium hydroxide combined with approximately 2,100 gm. of casein. 4. The equivalent combining weight of casein for this base is just half the minimal molecular weight as calculated from the sulfur and phosphorus content, and one-sixth the minimal molecular weight calculated from the tryptophane content of casein. 5. From the study of systems containing the protein and very small amounts of sodium hydroxide it was possible to determine the solubility of the casein molecule, and also the degree to which it dissociated as a divalent acid and combined with base. 6. Solubility in such systems increased in direct proportion to the amount of sodium hydroxide they contained. 7. The concentration of the soluble casein compound varied inversely as the square of the hydrogen ion concentration, directly as the solubility of the casein molecule, Su, and as the constants Ka₁ and Ka₂ defining its acid dissociation. 8. The product of the solubility of the casein molecule and its acid dissociation constants yields the solubility product constant, Su·Ka₁·Ka₂ = 2.2 x 10^–12 gm. casein per liter at 25°C. 9. The solubility of the casein molecule has been estimated from this constant, and also from the relation between the solubility of the casein and the sodium hydroxide concentration, to be approximately 0.09 gm. per liter at 25°C. 10. The product of the acid dissociation constants, Ka₁ and Ka₂, must therefore be 24 x 10^–12N. 11. It is believed that these constants completely characterize the solubility of casein in systems containing the protein and small amounts of sodium hydroxide.     

腹泻小儿便中双歧杆菌含量变化和红细胞免疫功能关系的探讨

目的：研究腹泻小儿便中双歧杆菌含量变化与红细胞免疫的关系。方法：对42例腹泻小儿和38例正常小儿进行了粪便双歧杆菌含量、红细胞CR1数量及红细胞自然粘附肿瘤花环率（NTRR）的测定。结果：与正常小儿相比,腹泻小儿粪便中双歧杆菌含量明显下降,红细胞CR1数量无明显改变,NTRR明显下降。结论：小儿肠道内双歧杆菌含量与其红细胞CR1免疫活性有相关关系。     

PROTEIN COAGULATION AND ITS REVERSAL : THE REVERSAL OF THE COAGULATION OF HEMOGLOBIN

1. The preparation from completely coagulated hemoglobin of crystalline soluble hemoglobin is described. 2. This soluble hemoglobin by all the tests tried has been indistinguishable from normal native hemoglobin which has never been coagulated. 3. The coagulation of hemoglobin is probably reversible. 4. Since hemoglobin is a typical coagulable protein, protein coagulation in general is probably reversible.     

晶状体生化的研究——亚硒酸钠诱发大鼠白内障晶状体中巯基、二硫键及维生素C的含量

我们测定了正常及亚硒酸钠诱发的白内障大鼠晶状体中非蛋白质巯基、蛋白质巯基、蛋白质结合巯基和维生素C的含量,发现随着白内障的进展非蛋白质巯基及蛋白质巯基均减少,蛋白质结合巯基在核混浊时增加,而在整个晶状体混浊时下降到与正常对照组相近,在白内障形成过程中二硫交联的蛋白质含量明显增加,而维生素C含量似乎无明显变化。     

l-THREONINE DEAMINASE IN MARINE PLANKTONIC ALGAE. II. DISULFIDE AND SULFHYDRYL GROUP REQUIREMENTS OF ENZYME ACTIVITY IN TWO CRYPTOPHYTES1

The “biosynthetic”l threonine (deaminating) dehydratase of 2 cryptophytes (Chroomonas salina and Hemiselmis virescens) showed sensitive inhibition from all thiols tested (dithiothreitol, cysteine, etc.) but no effect from ascorbic acid or reduced NAD. By contrast, the enzyme activities from 5 noncryptophyceaen unicellular algae (2 cyanophytes, 1 rhodophyte, 1 diatom, 1 chlorophyte) were generally not affected by any of these reagents. The thiol reagent inhibition of the cryptophyte enzymes (1) achieved saturation with 60–70% reduction in activity, (2) was considerably reduced by pretreatment of the enzymes with l -threonine and l -isoleucine, and (3) was partially reversed by subsequent treatment with arsenite and exposure to air. It was deduced that such inhibitions were caused by thiol-specific reduction of enzyme-protein disulfide groups essential for the full expression of activity and that these groups were susceptible to ready reductive cleavage and oxidative restoration. This disulfide requirement, unique to the cryptophytes, may be the first recorded case of such a property of threonine dehydratase from all forms of life hitherto studied. The additional activity requirement of the cryptophyte enzymes for sulfhydryl groups (which requirement was common to all the algal enzymes) was confirmed (1) by the study of their sensitivity to inhibition from mercurials and disulfide-sulfhydryl exchanging reagents, and (2) by the partial reversal of these inhibitions from subsequent treatment with dithio-threitol. Both cryptophyte enzymes were densitized to feedback inhibition from l -isoleucine by prior exposure to subinhibitory concentrations of HgCl₂ or dithiodipyridine.     

五步蛇主要数量性状与排毒量的相关分析

本文首次报道了五步蛇身体主要部位的数量性状与其排毒量的关系。其中我们发现与排毒量有关的部位是头积、体粗、体重、佛指长。     

STUDIES IN THE PHYSICAL CHEMISTRY OF THE PROTEINS : III. THE RELATION BETWEEN THE AMINO ACID COMPOSITION OF CASEIN AND ITS CAPACITY TO COMBINE WITH BASE.

1. The methods of measuring the base-combining capacities of proteins have been considered, and the constants and corrections that are employed in their calculation have been critically examined. 2. The base-combining capacities of ten casein preparations have been determined. These differed from each other to a far greater extent than can be attributed to the experimental errors involved in their measurement and calculation. The variations were, moreover, systematic in manner, and can be explained as dependent upon the method employed in the preparation of the casein. 3. Casein that had never been exposed to greater alkalinities than those in which it exists in nature combined with approximately 0.0014 mols of sodium hydroxide per gm., while casein prepared nach Hammarsten, and casein that was saturated with base during its preparation, combined with approximately 0.0018 mols of sodium hydroxide per gm. 4. 1 mol of sodium hydroxide, therefore, combined with 735 gm. of casein that had not previously been exposed to alkaline reactions, or with 535 gm. of casein that had previously been saturated with base. 5. If the minimal molecular weight of casein, based upon its tryptophane content, is placed at 12,800, the native protein must, therefore, contain approximately eighteen acid groups, and in addition six acid groups that are released in alkaline solutions, and presumably represent internally bound groups. The total base-combining capacity therefore represents that of a substance with a molecular weight of 12,800 and containing twenty-four acid valences. 6. This base-combining capacity is no greater than can be accounted for on the basis of our knowledge of the structure and composition of casein. On the basis of a molecular weight of 12,800 casein contains at least 19 molecules of glutamic acid, 4 of aspartic, and 8 of hydroxyglutamic acid. If the amino acids in the protein molecule are bound to each other in polypeptide linkage, each of these thirty-one dicarboxylic acids should yield terminal groups. The ammonia in casein suggests that twelve of these groups are bound as amides. As many as nineteen carboxyl groups may, therefore, be free in the protein molecule. 7. Casein contains phosphorus. If this phosphorus represents phosphoric acid, and if we consider that all of the valences of this acid are either themselves free, or that they have liberated carboxyl groups by entering into the structure of the protein molecule, casein should contain nine additional acid groups. 8. Recent analytical results, therefore, indicate that casein contains at least nineteen, and possibly twenty-eight, free acid groups. The physicochemical measurements presented suggest that casein combines with base as though it contained twenty-four acid groups, of which six, or one-fourth, appear to be bound in the native protein. These experimental results are therefore in close agreement with the expectation on the basis of the classical theory of protein structure.     

夏谷籽粒蛋白质积累和旗叶蛋白质周转的关系

谷子灌浆期旗叶蛋白水解酶活性、籽粒和旗叶游离氨基酸含量均呈双峰曲线变化。第一峰在花后１３天;第二峰分别在２５、２８天。籽粒和旗叶游离氨基酸峰稍滞后,且后者不明显。籽粒总蛋白积累和干重增加均为Ｓ型变化,与旗叶总蛋白和可溶性蛋白变化趋势相反。花后２２─２５天上述物质和酶活变化均出现─转折。转折点之前,蛋白质合成和周转活跃;之后,由叶片蛋白酶活性升高引起的旗叶氮素撤离对籽粒蛋白质积累仍有一定贡献（约１０％）。     

蚤类在宿主体表的分布及温度和蚤数的关系

本文研究了方形黄鼠蚤松江亚种Citellophilus tesquorum sungaris和二齿新蚤Neopsylla bidentatiformis在小白鼠体表的分布及环境温度和体表蚤数的关系.结果证明,在各种温度下,鼠背部蚤数均多于腹部.5℃时,鼠前部蚤数多于后部;10—20℃时,前部与后部蚤数相近;25—40℃时,后部蚤数多于前部.鼠后背部蚤数随温度升高而增多,前腹部蚤数随温度升高而减少;温度越高二者之差越大,说明温度越高蚤越向后背部集中.鼠体蚤数较少时,背部与腹部,前部与后部,蚤数差别明显;鼠体蚤数过多时,各部位蚤数无大差别.     

THE RELATIONSHIP BETWEEN DIURNAL VARIATION OF THE NUMBER OF CELLS IN MITOSIS AND OF THE NUMBER OF CELLS SYNTHESIZING DNA IN THE EPITHELIUM OF THE HAMSTER CHEEK POUCH

1. The numbers of cells in mitosis and in DNA synthesis in the epithelium of the hamster cheek pouch have been studied at different times of the day and night. 2. By accumulation of mitotic cells using colcemid, both the rate of entry of cells into mitosis and the duration of mitosis have been estimated at two different times of day. 3. A diurnal variation has been demonstrated in both the mitotic index and in the tritiated thymidine labelling index. Although these variations are of different amplitude and timing, the experimental data fit closely to the hypothesis that the diurnal mitotic variation is the result of a partially synchronous population moving through the DNA synthetic period. No direct action on the mitotic process need be postulated. 4. From the results of mitotic accumulation, it is clear that the rate of entry of cells into mitosis depends on the time of day at which this is studied. There is also the possibility that the duration of mitosis is slightly longer when the mitotic index is high. 5. It is concluded that, at least in the epithelium of the hamster cheek pouch, the diurnal rhythm in the number of mitoses present is a reflection of the diurnal variation in the number of cells synthesizing DNA at some time earlier. Small fluctuations in the mitotic pattern imposed by this partially synchronous population moving from S into mitosis, could be caused by slight variations in the duration of mitosis.     

AUXIN-INDUCED GROWTH OF TUBER TISSUE OF JERUSALEM ARTICHOKE: II. THE RELATION TO PROTEIN AND NUCLEIC ACID METABOLISM

The following results were obtained using tissue slices excisedfrom cold-stored Jerusalem artichoke tuber. 1. Increase in protein content of the tissue was small duringthe washing (i.e. "aging"), and great in the growth phase, particularlyin washed tissue. 2. RNA content of tissue increased during the growth periodsimilarly in non-growing tissue (in water) and actively growingtissue (in 2,4-D plus KIN). 3. Both RNA and DNA increased during the washing, the increasebeing greater in RNA than in DNA. This RNA increase was enhancedby gibberellic acid. 4. 2-Thiouracil, 8-azaguanine, puromycin, and mitomycin C givenat the washing inhibited the subsequent growth. The effect ofthese inhibitors was not significant when they were given inthe growth period. 5. Mitomycin C reduced the basophilia of nuclei and made themswell, as did deoxyribonuclease. 6. The effect of inhibitors of nucleic acid metabolism was reversedto some extent by gibberellic acid and by kinetin. 7. Chloramphenicol inhibited the growth strongly if given inthe growing period, but not so strongly if given during thewashing. 8. An autoradiographic study using ³H-cytidine suggested thatRNA is synthesized in nucleus during the period of washing andis transferred to cytoplasm via nucleolus. It is conjectured that the RNA synthesized during the agingis responsible for the expansion growth to be caused later byauxin or auxin plus kinetin. (Received September 4, 1965; )