Determination of Melting Sequences in DNA and DNA-Protein Complexes by Difference Spectra

A graphical formula is presented for determining the base ratio of melted DNA. By use of this formula, the composition of sequences which melt in different portions of the melting curves of Clostridium DNA, Escherichia coli DNA, and mouse DNA were determined. As the DNA melts, the per cent of adenine and thymine (AT) in the melted sequences decreases linearly with temperature. The average composition of sequences which melt in a given part of the melting curve is proportional to the base ratio of the DNA. The concentration and average composition of sequences were determined for three parts of the melting curves of the DNA samples, and a frequency distribution curve was constructed. The curve is symmetrical and has a maximum at about 56% AT. The distribution of GC-rich sequences on the E. coli chromosome was estimated by shearing, partially melting, and fractionating the DNA on hydroxylapatite. GC-rich sequences appear to occur every thousand base pairs, and have a maximum length of about 180 base pairs. The graphical formula was applied to the determination of the composition of sequences which melt in different parts of the melting curve of chromatin. Throughout the melting curve, the composition of the melting sequences is about 60% AT, which appears to suggest that relatively long sequences are melting simultaneously. Their melting temperature may be a function of the composition of the protein on different parts of the DNA. The problem of light scattering in DNA-protein and DNA was also investigated. A formula is presented which corrects for light scattering by relating the intensity of the scattered light to the rate of change of absorbance of DNA with wavelength.     

Methods for Detecting Food-borne Enteroviruses

A method previously reported for detecting virus in a model system composed of cottage cheese contaminated with coxsackievirus type A9 has been adapted to detecting selected strains of enteroviruses in a variety of foods. Bentonite is omitted and serum is added for extracting virus from low-protein foods. Samples of foods, usually 25 g, must contain at least 3 to 4 plaque-forming units for a 50% probability of detecting virus. Sensitivity in detecting echovirus type 6 was lower than that for the other viruses used. After extraction from potato salad, poliovirus type 2 was completely reactivated if it had been neutralized with coproantibody, but it was only partially reactivated if neutralized with hyperimmune rabbit serum.     

Der Einfluß des Zeugungsalters auf die Mutationen zu Hämophilie A

Assuming that in sibships with sporadic hemophilia the mothers of the patients are already carriers the relationship between the age of the mother's father at birth of the mother and frequency of hemophilia among his grandsons was examined.The ages of the mother's father at birth of the mother of 40 patients with sporadic hemophilia A was compared with that of a control-group as well as with the ages of the mother's fathers at birth of the mothers of patients from families where hemophilia A is inherited. The mean age of these grandfathers was found to be increased. Using the Mann-Whitney-U-test for comparing the ages of grandfathers of sporadic cases with the control-group there is to be found P=.0025, which is highly significant statistically. Comparing data from sibships with sporadic hemophilia with data from sibships where hemophilia is inherited there is no significant difference — perhaps according to the small number of inherited cases (19, P=0.065) —, but the deviation is in the same direction. Comparison of data both from the inherited cases and the control-group with data of sporadic cases gives P=.0087.There is perhaps a connection between parental age and number of children, but it is shown to have no important influence in our material. On the other hand fathers with a higher number of children are significant more frequent among the grandfathers than among the controls. This difference cannot be explained sufficiently. Between cases of sporadic and inherited hemophilia there is no clear cut difference.Certainly there exists a relationship between parental age and birth-rank. Therefore the mothers of sporadic cases — unlike to carriers of sibships with inherited hemophilia — take clearly higher ranks in birth-order than it is theoretically to be expected. Penrose published a method of separating the relative aetiological effects of birth-order and parental age. Using this method an influence of birth-order cannot be found after excluding the influence of parental age. Hence, paternal age seems to be the determining factor.It is discussed which model of mutation the hemophilia-mutation belongs to because of this relationship. We would count it to No. 2 of the classification given by Vogel where mutations are due to copy errors. So cases of sporadic hemophilia seem equal to those of sporadic achondroplasia.

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Diese Arbeit wurde der Medizinischen Fakultät der Universität Hamburg als Inaugural-dissertation vorgelegt.

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Herrn Prof. Dr. F. Vogel danke ich für wertvolle Hinweise, insbesondere zur statistischen Methodik.     

BIOSYNTHESIS OF COLLAGEN AND NON-COLLAGEN PROTEIN DURING DEVELOPMENT OF THE CHICK CORNEA

The relationship between the rates of increase of corneal protein fractions and incorporation of labeled precursors has been examined during embryonic and early posthatching development of the chick corneal stroma. Non-collagen protein increased gradually from 9 through 20 days of incubation. Collagen accumulated approximately logarithmically through the 19th day, the most rapid rate occurring between 13 and 20 days of incubation. The rates at which labeled amino acids are incorporated into collagen in vivo and in vitro undergo marked changes during the last week of embryonic development, corresponding closely to the rate of collagen accumulation in vivo; whereas incorporation into non-collagen protein changes much less markedly. Changes in the rate of incorporation of precursors into collagen are not due to changes in the rate of conversion of collagen from the soluble to insoluble form, or to changes in the endogenous amino acid pool size. Chick embryo corneal stroma collagen turns over very slowly, if at all. Non-collagen protein turns over more rapidly. An increase in cell number, as indicated by DNA content, does not account for the increased rate of collagen synthesis between the 9th and 16th day of incubation. It is concluded that the observed changes in collagen synthesis reflect changing activities in the individual cornea fibroblasts. These activities are comparable in the intact tissue in vivo and in isolated corneas in vitro.     

Histochemische Untersuchungen an der Leber nach Verbrennung

Zusammenfassung An männlichen weißen Ratten wurde das Verhalten des Enzymmusters der Leber 72 Std nach Verbrühung untersucht. Es fanden sich Veränderungen im Sinne einer Aktivitätsverminderung bei der NAD-Cytochrom-c-Reduktase, Isocitronensäuredehydrogenase, ATPase und den PJS-positiven Substanzen. Eine Zunahme der Aktivität wurde bei der alkalischen Phosphatase und einigen Esterasen beobachtet, ebenso traten auch die scharlachrotfärbbaren Substanzen auf. Uneinheitliche Reaktionen zeigten die Succinodehydrogenase sowie die Leucinaminopeptidase, unverändert blieben Glukose-6-phosphat-dehydrogenase, -Glucuronidase, saure Phosphatase, die o-Diacetylbenzol- und die DDD-Reaktion.     

HIV-gp120 can block CD4-class II MHC-mediated adhesion

A possible component of the immune dysfunction associated with infection by HIV is the inhibition of CD4 function resulting from the avid binding of soluble HIV envelope glycoprotein (gp120) to cell surface CD4. We assessed CD4 function by measuring the ability of CD4+ T cells to form conjugates with cell size lipid vesicles, artificial target cells (ATC), bearing the natural ligand for CD4, MHC class II proteins. Conjugate formation was a transient process with the greatest number of specific cell to ATC conjugates found after approximately 30 min of incubation at 37 degrees C. Addition of gp120 specifically blocked conjugates between CD4+ cells and class II ATC in a concentration-dependent manner. These data indicate that T lymphocyte adhesion mediated by CD4 is a dynamic event and that binding of gp120 to CD4 is able to disrupt the normal progression of the interaction between CD4+ T lymphocytes and class II+ APC.     

Inversion of gravitropism by symmetric blue light on the clinostat

Gravitropic stimulation of maize (Zea mays L.) seedlings resulted in a continuous curvature of the coleoptiles in a direction opposing the vector of gravity when the seedlings were rotated on a horizontal clinostat. The orientation of this response, however, was reversed when the gravitropic stimulation was preceeded by symmetric preirradiation with blue light (12.7 mol photons·m^–2). The fluence-response curve of this blue light exhibited a lower threshold at 0.5 mol·m^–2, and could be separated into two parts: fluences exceeding 5 mol·m^–2 reversed the direction of the gravitropic response, whereas for a range between the threshold and 4 mol·m^–2 a split population was obtained. In all cases a very strong curvature resulted either in the direction of gravity or in the opposite orientation. A minor fraction of seedlings, however, curved towards the caryopsis. Furthermore, the capacity of blue light to reverse the direction of the gravitropic response disappeared with the duration of gravitropic stimulation and it depended on the delay time between both stimulations. Thistonic blue-light influence appears to be transient, which is in contrast to the stability observed fortropistic blue-light effects.This work was supported by the Deutsche Forschungsgemeinschaft.     

Characteristics and N-terminal amino acid sequence of a manganese peroxidase purified from Lentinula edodes cultures grown on a commercial wood substrate

Summary Extracellular culture filtrates from ligninolytic cultures of the lignin-degrading basidiomycete Lentinula (syn. Lentinus) edodes (Berk.) Pegler contained one major peroxidase when grown on a commercial oak-wood substrate. The peroxidase was purified by polyethylenimine clarification, anion-exchange chromatography, and hydrophobic-interaction HPLC. The enzyme (MnP1) was a heme-iron protein with an apparent molecular weight of 44 600 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels and an isoelectric point of pH 3.2. The native enzyme had an absorption maximum at 407 nm, which shifted to 420 nm upon H₂O₂ addition. The pyridine-hemochrome-absorption spectrum indicated that one heme group was present per enzyme as protoporphyrin IX. N-terminal amino acid sequencing showed that MnP1 had higher sequence homology with manganese peroxidases than with lignin peroxidases reported from Phanerochaete chrysosporium. L. edodes MnP1 was capable of oxidizing lignin and lignin-model compounds in the presence of manganese and H₂O₂.On leave from the Department of Biochemistry, University of Otago, P. O. Box 56, Dunedin, New Zealand.Research carried out while a visiting scientist at the USDA Forest Products Laboratory from the National Chemistry Laboratory, Pune, India 41 1008 Offprint requests to: I. T. Forrester