Isoenzyme patterns of human liver alpha-L-fucosidase during development.

The pattern of human liver α-l-fucosidase isoenzymes during development has been studied by isoelectric focusing. Seven isoenzymes have been found in livers from fetuses, children and adults. The pattern of isoenzymes appears to change during development. Whereas the most neutral form (I) of α-l-fucosidase is prominent in early fetal development (105–109 days gestation), it is greatly diminished in amount later in fetal development (123–124 days gestation) and in children and adults. Incubation of adult human liver α-l-fucosidase with neuraminidase eliminates the five most acid forms of the enzyme and greatly increases the amount of the most neutral form (I). It is thus possible that sialylation of the most neutral form of α-l-fucosidase accounts, at least in part, for the changes in fucosidase isoenzyme patterns during development.     

Definitive relationships among chemical structure, carcinogenicity and mutagenicity for 301 chemicals tested by the U.S. NTP.

An analysis is presented in which are evaluated correlations among chemical structure, mutagenicity to Salmonella, and carcinogenicity to rats and mice among 301 chemicals tested by the U.S. NTP. Overall, there was a high correlation between structural alerts to DNA reactivity and mutagenicity, but the correlation of either property with carcinogenicity was low. If rodent carcinogenicity is regarded as a singular property of chemicals, then neither structural alerts nor mutagenicity to Salmonella are effective in its prediction. Given this, the database was fragmented and new correlations sought between the derived sub-groups. First, the 301 chemicals were segregated into six broad chemical groupings. Second, the rodent cancer data were partially segregated by target tissue. Using the previously assigned structural alerts to DNA reactivity (electrophilicity), the chemicals were split into 154 alerting chemicals and 147 non-alerting chemicals. The alerting chemicals were split into three chemical groups; aromatic amino/nitro-types, alkylating agents and miscellaneous structurally-alerting groups. The non-alerting chemicals were subjectively split into three broad categories; non-alerting, non-alerting containing a non-reactive halogen group, and non-alerting chemical with minor concerns about a possible structural alert. The tumor data for all 301 chemicals are re-presented according to these six chemical groupings. The most significant findings to emerge from comparisons among these six groups of chemicals were as follows: (a) Most of the rodent carcinogens, including most of the 2-species and/or multiple site carcinogens, were among the structurally alerting chemicals. (b) Most of the structurally alerting chemicals were mutagenic; 84% of the carcinogens and 66% of the non-carcinogens. 100% of the 33 aromatic amino/nitro-type 2-species carcinogens were mutagenic. Thus, for structurally alerting chemicals, the Salmonella assay showed high sensitivity and low specificity (0.84 and 0.33, respectively). (c) Among the 147 non-alerting chemicals less than 5% were mutagenic, whether they were carcinogens or non-carcinogens (sensitivity 0.04).     

1H NMR studies of heme pocket conformation in zinc-substituted leghemoglobin, a diamagnetic analog of deoxyleghemoglobin

Reconstitution of apoleghemoglobin with zinc protoporphyrin IX is reported. NMR spectra show that the reconstitution is orientation specific and that there is no detectable heme isomerism or conformational heterogeneity. Resonances of heme substituents and distal and proximal amino acid protons have been assigned. Only minor differences in porphyrin-protein packing occur between zinc leghemoglobin and the CO complex of ferrous leghemoglobin. The zinc is five-coordinate and is ligated by the proximal histidine. Comparisons with diamagnetic six-coordinate complexes show that the distal His-61 and Leu-65 side chains move away from the binding site upon coordination of exogenous ligands. Conformational changes are minimal when the ligand is O2.     

Analysis of recombinant DNA clones of the endogenous BALB/c murine leukemia virus WN1802N: variation in long terminal repeat length

We analyzed 15 recombinant DNA clones of the unintegrated closed circular DNA intermediate of the BALB/c endogenous ecotropic murine leukemia virus WN1802N. Thirteen of these clones had an insert which corresponded to the complete murine leukemia virus genome. Of these, six contained a single long terminal repeat (LTR) and seven contained two LTRs. The viral genomes in nine clones had an LTR of 520 base pairs (bp), one had an LTR of 570 bp, three had an LTR of 600 bp, and one had an LTR of 670 bp. Restriction endonuclease analysis demonstrated that the size variability resides in the U3 region. Seven of eight clones which yielded infectious virus by DNA transfection had the 520-bp LTR, and the other had a 600-bp LTR. More detailed examination of plasmid subclones of three isolates with different-sized LTRs revealed that the approximate position which varies in the U3 region corresponds to the 72-bp repeat region of Moloney sarcoma virus. Possible consequences of these variations are discussed.     

Reversal of Fv-1 host range by in vitro restriction endonuclease fragment exchange between molecular clones of N-tropic and B-tropic murine leukemia virus genomes.

We molecularly cloned unintegrated viral DNA of the BALB/c endogenous N-tropic and B-tropic murine leukemia retroviruses and in vitro passaged N-tropic Gross (passage A) murine leukemia retroviruses. Recombinant genomes were constructed in vitro by exchanging homologous restriction enzyme fragments from N- or B-tropic parents and subsequent recloning. Infectious virus was recovered after transfection of these recombinant genomes into NIH-3T3 cells and cocultivation with the Fv-1 nonrestrictive SC-1 cells. XC plaque assays of recombinant virus progeny on Fv-ln and Fv-lb cells indicated that the Fv-l host range was determined by sequences located between the BamHI site in the p30 region of the gag gene (1.6 kilobase pairs from the left end of the map) and the HindIII site located in the pol gene (2.9 kilobase pairs from the left end of the map).     

Natural history of experimental woodchuck hepatitis virus infection: molecular virologic features of the pancreas, kidney, ovary, and testis.

The kinetic patterns of woodchuck hepatitis virus (WHV) infection were monitored in the pancreas, kidneys, ovaries, and testes. Groups of woodchucks experimentally infected with a standardized inoculum of WHV were sacrificed at different times over a 65-week period beginning in the preacute phase of viral infection and continuing to the period of serologic recovery or the establishment of chronic infections and subsequent hepatocellular carcinoma (B. E. Korba, P. J. Cote, F. V. Wells, B. Baldwin, H. Popper, R. H. Purcell, B. C. Tennant, and J. L. Gerin, J. Virol. 63:1360-1370, 1989). Tissues from an additional group of long-term (2 to 3 years) chronic WHV carriers which had been infected with the same WHV inocula were also examined. Viral DNA replication intermediates were found in all four tissues during the acute phase of WHV infection. However, WHV DNA replication intermediates were observed only in the kidneys of a small proportion of the chronically infected animals. Following the acute phase of infection, WHV DNA was present only in the pancreas, kidneys, and ovaries of the chronically infected woodchucks. A progressive evolution of different WHV genomic forms related to the replicative state of WHV was observed in these tissues. Histologic evaluation of these four tissues revealed only minimal, localized lesions which were not correlated with the state of WHV activity. The observations compiled in this study further extend the tissue tropism of WHV.     

Natural history of woodchuck hepatitis virus infections during the course of experimental viral infection: molecular virologic features of the liver and lymphoid tissues.

In this study, the kinetic patterns of woodchuck hepatitis virus (WHV) infection were monitored in the liver and the five primary components of the lymphoid system (peripheral blood lymphocytes, lymph nodes, bone marrow, spleen, and thymus). Groups of woodchucks experimentally infected with a standardized inoculum of WHV were sacrificed at different times over a 65-week period beginning in the preacute phase of viral infection and continuing to the period of serologic recovery or the establishment of chronic infections and subsequent hepatocellular carcinoma. Infection by WHV was not limited to the liver but involved the major components of the lymphoid system during all stages of virus infection. A complex series of kinetic patterns was observed for the appearance of WHV DNA in the different lymphoid compartments and the liver during the entire course of viral infection. A progressive evolution of different WHV genomic forms related to the replicative state of WHV was also observed. Lymphoid cells of the bone marrow were the first cells in which WHV DNA was detected, followed in order by the liver, the spleen, peripheral blood lymphocytes, lymph nodes, and finally the thymus. Several differences were observed in the cellular WHV DNA patterns between woodchucks that developed chronic WHV infections and those that serologically recovered from acute WHV infections. The observations compiled in this study indicate that the host lymphoid system is intimately involved in the natural history of hepadnavirus infections from the earliest stages of virus entry.