Proteinchemical characterization of three structurally distinct domains along the protofilament unit of desmin 10 nm filaments

Limited chymotryptic cleavage of soluble chicken gizzard desmin protofilaments allows the characterization of three structurally distinct domains. A surface-exposed very basic amino-terminal region (the headpiece) with an amino acid sequence excluding a-helical organization (7.5 kd) is separated from the perhaps globular carboxy-terminal 48 residues (the tailpiece) by a distinctly different middle domain of approximately 330 residues. This 38 kd domain is very rich in α-helix (at least 83%), and electron microscopy reveals a thin rod with a length of 500 ± 50 Å. Amino acid sequence data also show that the rod domain is interrupted by a nonhelical portion. An a-helical array is able to form a coiled-coil spanning the carboxy-terminal half of the 38 kd domain. The a-type diffraction pattern of 10 nm filaments arises from a coiled-coil conformation displayed through most but not all of the middle domain of the protofilaments.     

Determination and postnatal development of fructokinase activity in swine liver]

Starting from the spectrophotometric method, described in ADELMAN et al., optimal reaction conditions for the measurement of fructokinase (ketohexokinase) in pig liver were systematically studied. It was necessary to increase the concentration of the substrate and further to lower the concentration of ATP for an optimal Mg: ATP-radio of 2:1. Using the optimized method fructokinase activity was determined in pig liver in relation to age, beginning from the last days of pregnancy to puberty. In liver of fetuses and newborn piglets during the first two days of life no or only a minute activity of the fructokinase was recorded. Therefore, the high level of fructose in fetal blood results from the inability of the fetus to metabolize fructose synthezised in the placenta or the fetal organs. At the end of the first week of life the activity of fructokinase was 10 times, after the second week 15-20 times higher than at birth. This high level remains constant during the suckling period and after weaning. For this reason, piglets after the first week of life are able to metabolize fructose and after the third week to form glucose and to release it into circulation. In adult pigs the activity of fructokinase in the liver decreases slightly. It corresponds-as in rat and human-to the elimination rate of experimentally applied fructose from the circulation. Therefore, this enzyme even in pigs is of significant importance for the utilization of fructose.     

Increase in liver and kidney deoxycytidine kinase activity linked to neoplastic transformation

Deoxycytidine kinase activity in normal rat liver cytosol was low (0.8 nmol/hr/mg protein); it increased 2–26-fold in 12 lines of chemically-induced, transplantable rat hepatomas of different growth rates. The increased kinase activity correlated positively with the hepatoma growth rate. The kinase activity did not change in the regenerating liver and the activity in the differentiating, neonatal rat liver was similar to values in adult liver. Deoxycytidine kinase activity in 2 chemically-induced, transplantable rat kidney tumors was increased to twice the value found in normal renal cortex. Among 15 normal rat tissues examined the highest kinase activities were observed in thymus, bone marrow and spleen. Of the normal and malignant rat tissues tested, only testis had detectable cytidine deaminase activity.     

Amino acid transport in normal and Rous sarcoma virus-transformed chicken embryo fibroblasts.

A study was made of the transport of a variety of amino acids by uninfected and Rous sarcoma virus-infected chicken embryo fibroblasts. Following a period of amino acid starvation, transformed, but not normal cells, showed increased levels of transport for alpha-aminoisobutyric acid, proline and alanine, three amino acids which are transported primarily by the A transport system. There was no starvation-induced increase in the transport of leucine, phenylalanine, lysine, or cycloleucine. In the absence of starvation, normal and transformed cells exhibited comparable rates of amino acid transport. Cycloheximide was able to block the increase in uptake. The enhanced uptake was characterized by an increase in Vmax for transport and little change in Km. The data demonstrate that an alteration in the regulation of the A amino acid transport system is an early event in malignant transformation by Rous sarcoma virus. However, since this alteration in made manifest only following a period of starvation, our findings suggest that increased amino acid uptake does not play a role in generating the other manifestations of the transformed state seen in cell culture.     

The formation of dipeptides from amino acids and the 2′(3′)-glycyl ester of an adenylate

Summary The yields of dipeptide obtained from the reaction of 0.2M 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) and 0.2M amino acid at pH 8.2 ranged from 0.1% to 35.5% for a group of 15 amino acids. The yields of glyser (35.3%), gly-cys (11.8%) and gly-thr (5.4%) were considerably greater than dipeptide yields obtained from any of the other 12 amino acids ( 1.7%). Aminolysis of 0.05M 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) by 0.4M serine ethyl ester yielded 53% glycylserine diketopiperazine, via N-(glycyl)-serine ethyl ester as a transient intermediate. The prebiotic significance of these reactions is discussed.Abbreviations MepA adenosine-5-(O-methylphosphate) - MepA-gly 2(3)-O-(glycyl)-adenosine-5-(O-methylphosphate) - DKP diketopiperazine - serOEt serine ethyl ester - gly-serOEt N-(glycyl)-serine ethyl ester - Boc-gly N-tertbutyloxycarbonylglycine - cyclo-(gly-ser-) glycylserine diketo-piperazine - O-gly-ser O-glycylserine - O-(gly)-gly-ser O-(glycyl)-glycylserine - gly-ser N-glycylserine     

The distribution of keratin type intermediate filaments in human breast cancer. An immunohistological study

Antibodies to different intermediate filament proteins can be used to distinguish cells of epithelial, mesenchymal, muscle, glial and neuronal origin. Antibodies to prekeratin which characterize cells of epithelial origin, and antibodies to vimentin which recognize cells of mesenchymal origin have been used to study twenty cases of breast carcinoma (sixteen infiltrating ductal carcinomas and four infiltrating intraductal carcinomas), two cases of cystic breast disease, two fibroadenomas and one case of benign cystosarcoma phylloides. The prekeratin and vimentin were detected using specific antibodies to these proteins by immunofluorescence microscopy using alcohol fixed paraffin-embedded tissues. In eighteen out of the twenty carcinomas the tumor cells were strongly and specifically stained by antibodies to prekeratin. DIfferent tumors gave different patterns of prekeratin staining. In contrast, when the same specimens were tested with the vimentin antibody, the tumor cells were unstained, and instead only the usual strong staining to fibroblasts and blood vessels in the stroma was observed. In cystic breast disease, fibroadenomas, and benign cystosarcoma phylloides, cells of epithelial origin were strongly stained by the prekeratin but not by the vimentin antibody.