Interaction of mitochondrial malate dehydrogenase monomer with phospholipid vesicles.

The association between bovine and porcine mitochondrial malate dehydrogenase (EC 1.1.1.37) and phospholipid vesicles was investigated. At concentrations at which malate dehydrogenase exists as a dimer, entrapment within the aqueous compartment but not binding of the 14C-labelled enzyme was observed. The dissociated enzyme was labile to moderate heat and to p-chloromercuribenzoate, but in both cases inactivation was decreased by incubation with suspensions of charged phospholipid vesicles. This suggested an interaction between enzyme subunits and phospholipid, and this was confirmed by direct binding measurements and by studies that followed changes in the fluorescein-labelled enzyme. The circular-dichroism spectra of the enzyme indicated a high alpha-helix content, and suggested that a small conformational change occurred when the enzyme dissociated. Fluorescence data also suggested less-rigid molecules after dissociation. A possible mechanism, based on the flexibility of enzyme monomer and its interaction with phospholipids, by which mitochondrial matrix enzymes are specifically localized in cells, is discussed.     

Chemical and physical properties of mammalian mitochondrial aminoacyl-transfer RNAs. I. Molecular weights of mitochondrial leucyl- and methionyl-transfer RNAs.

The sedimentation and electrophoretic properties of Syrian hamster cytosolix and mitochondrial methionyl- and leucyl- +RNAs have been compared under denaturing conditions. Mitochondrial leucyl-tRNA could be separated into three species by chromatography on RPC-5. Their apparent molecule weights as determined by polyacrylamide slab gel elecltrophoresis were 23 000 for one species and 24 000 for the other two compared to the five cytosolic leucyl-tRNA species whose apparent molecular weights ranged from 26 000 to 28 000. Mitochondrial leucyl-tRNAs sedimented more slowly than their cytosolic counterparts, again indicating a lower molecular weight. The apparent molecular weights of the mitochondrial methionyl-tRNAs were identical or only slightly lower than their cytosolic counterparts as determined by polyacrylamide slab gel electrophoresis but both mitochondrial methionyl-tRNA and formylmethionyl-tRNA sedimented slightly more slowly than cytolsolic methionyl-tRNA. It is suggested that mitochondrial tRNAs fall into the size range of other t RNAs and might be uniform in size.     

Chemical and physical properties of mammalian mitochondrial aminoacyl-transfer RNAs. II. Analysis of 7-methylguanosine in mitochondrial and cytosolic aminoacyl-transfer RNAs.

The 7-methylguanosine (m7G) content of two individual mitochondrial tRNAs, labelled in the aminoacyl moiety was assayed by the specific cleavage of the tRNA at this nucleotide followed by electrophoretic analysis to identify the 3'-terminal fragment of the tRNA. Neither Syriam hamster mitochondrial tRNALeu nor tRNAMet were found to contain m7G. In contrast, cytosolic tRNAMetS were cleaved indicating the presence of m7G, apparently 27--28 and 29 nucleotides from their 3' terminus. Cystolic tRNALeu was not cleaved. These results are discussed in relationship to the reported low content of methylated nucleosides in mitochondrial 4 S RNA.     

Metabolites of estradiol-17β in guinea pig uterus late in pregnancy

The metabolism of estradiol-17β by the guinea pig uterus late in pregnancy was studied $\text{in vivo}$ and $\text{in vitro}$.Whole uteri were examined for estrogen metabolites one hour following an intravenous injection of [³H]-estradiol-17β or uterine sections were examined after incubation for one hour at 37°C in medium containing [³H]-estradiol-17β.In both instances uterine tissue metabolized estradiol-17g to five products: estrone, estrone-3-sulfate, 17β-estradiol-3-sulfate, estrone-3-glucuronide and 17β-estradiol-3-glucuronide. Of the total radioactive products 11 – 43% were glucuronides, 17 – 26% were sulfates and 4 – 17% was estrone. These results indicate that the guinea pig uterus actively transforms estradiol-17β into glucuronides and sulfates late in pregnancy.     

The cytogenetic systems of grasshoppers and locusts

The three endemic species of the Australian genusTolgadia are all distributed in the northern tropical and subtropical areas. They are distinguishable both on morphological and cytological grounds. All three have a derived neo XY, neo XX sex chromosome system involving distinct elements of a basically 11-membered autosome set, namely (spacies-1) (bivittata) and (infirma). Additionallybivittata is homozygous for two autosomal fusions involving and giving 2n=18 as compared to the complement of 2n=22 present in the other two species. Two of the species,bivittata and species-1, have a comparable mean cell chiasma frequency despite their difference in chromosome number. That ofinfirma is significantly higher. Coupled with this, some of the populations ofinfirma are polymorphic for supernumerary heterochromatic segments, principally on the smallest autosome. Such segments increase the mean cell chiasma frequency still further. Thusinfirma, which is the least habitat restricted despite its brachypterous nature, not only has the highest mean cell chiasma frequency but, in addition, has at its disposal a polymorphism capable of magnifying this difference.     

Growth and characterization of human skin epithelial cell cultures

Summary In 129 of 140 attempts, human skin cells were successfully cultured on the dermal collagen bed of sterile, dead pigskin. Diploid epithelial cells grew selectively on the collagen bed; fibroblasts grew on the glass surfaces of the culture dishes. The cultures could be subdivided physically up to six times at a 1:2 split ratio, but at least 24 to 48 cell generations were produced over the months the cells could be carried. Much of the cell multiplication resulted in maturation into distinct basal, squamous, granular, and keratinized cell layers. The cultured cells were considered epithelial because of their shape, possession of intercellular bridges, desmosomes and tonofibrils, and because they formed maturating epithelium in vitro and upon transplantation back to the original human donor. As the cells grew they digested the pigskin collagen, thus producing clear zones that could be used to monitor and quantitate cell growth. Multiplication of epithelial cells, rather than migration, was indicated by mitotic figures in colchicine-treated cultures and by DNA synthesis. Expert technical assistance was provided by Nancy Allen (cell culture); William Towler (electron microscopy); James Malone, Nona Scaife, and Joy M. Nicolet (cytogenetics); R. Thomas Campbell and Dorothy Sarver (photography); and V. L. Angerstein, Susan Ekker, and Arnater Yarbrough (histology). This work was supported by The United Fund Cancer Society of Summit County, the Greater Cleveland Associated Foundation (grant no. 3G3490X1), the National Institute of General Medical Services (grant no. 1 R01 GM 21929-01), and the Charles E. Merrill Trust.     

Synthesis of mammalian mitochondrial rRNA at low temperature

L cells incubated at 19 °C synthesize mitochondrial rRNA but not cytosol rRNA. In addition to 16 and 13S mitochondrial rRNA, mitochondrial RNA sedimenting at higher S values was also synthesized. The processing of mitochondrial rRNA is slower at 19 °C than at 37 °C.     

Causes and consequences of Robertsonian exchange

At least two types of Robertsonian exchange are now known in the acrocentric chromosomes of man. Both types involve breakage in the arms adjacent to the centromere. Evidence is presented for a third type of exchange, one involving breakage within the centromere itself, in the grasshopper Percassa rugifrons. In this species, which is regularly homozygous for a single fusion metacentric, the eighteen rod autosomes have small but pronounced granules at the centric end of the chromosome. When C-banded these granules show differential Giemsa staining and appear to represent centromeric chromomeres; these chromomeres are lacking in the metacentric fusion product. Equivalent fusions may have occurred in some mammal species too and possible examples of this are discussed in sheep and mice. The Percassa fusion has led to a modification in both the frequency and the distribution of chiasmata as judged by a comparison of these properties in the metacentric relative to the two next smallest rod equivalents. Comparable modifications are known to occur in other naturally occurring fusions but these changes are certainly not automatic consequences of fusion since they are not shown in at least some newly produced fusion mutants.     

An NMR investigation of electron transfer in the copper-protein, plastocyanin.

1. The proton NMR spectra of oxidised and reduced French bean plastocyanin have been recorded on a 270 MHz pulsed spctrometer. 2. The spectrum of a mixture containing the protein in the paramagnetic Cu(II) and diamagnetic Cu(I) states is a superposition of the separate spectra. When ferrirate spectra. 3. The results show that self-exchange between Cu(II)- and Cu(I)-plastocyanin is slow on the NMR time scale (kex less than 2-10(4) M-1-s-1 at 50 degrees C), and that electron transfer in the presence of ferricyanide is rapid (k greater than 1-10(5) M-1-s-1).