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2.  Nitrogen fixation specific regulatory genes of Klebsiella pneumoniae and Rhizobium meliloti share homology with the general nitrogen regulatory gene ntrC of K. pneumoniae. 
Nucleic Acids Research  1985;13(12):4539-4555.
We have determined the complete nucleotide sequences of three functionally related nitrogen assimilation regulatory genes from Klebsiella pneumoniae and Rhizobium meliloti. These genes are: 1) The K. pneumoniae general nitrogen assimilation regulatory gene ntrC (formerly called glnG), 2) the K. pneumoniae nif-specific regulatory gene nifA, and 3) an R. meliloti nif-specific regulatory gene that appears to be functionally analogous to the K. pneumoniae nifA gene. In addition to the DNA sequence data, gel-purified K. pneumoniae nifA protein was used to determine the amino acid composition of the nifA protein. The K. pneumoniae ntrC and nifA genes code for proteins of 52,259 and 53,319 d respectively. The R. meliloti nifA gene codes for a 59,968 d protein. A central region within each polypeptide, consisting of approximately 200 amino acids, is between 52% and 58% conserved among the three proteins. Neither the amino termini nor the carboxy termini show any conserved sequences. Together with data that shows that the three regulatory proteins activate promoters that share a common consensus sequence in the -10 (5'-TTGCA-3') and -23 (5'-CTGG-3') regions, the sequence data presented here suggest a common evolutionary origin for the three regulatory genes.
PMCID: PMC321805  PMID: 2989799
5.  Ro 32-3555, an orally active collagenase inhibitor, prevents cartilage breakdown in vitro and in vivo 
British Journal of Pharmacology  1997;121(3):540-546.
Ro 32-3555 (3(R)-(cyclopentylmethyl) - 2(R) - [(3,4,4 - trimethyl-2,5-dioxo-1-imidazolidinyl)methyl] - 4-oxo-4-piperidinobutyrohydroxamic acid) is a potent, competitive inhibitor of human collagenases 1, 2 and 3 (Ki values of 3.0, 4.4 and 3.4 nM, respectively). The compound is a selective inhibitor of collagenases over the related human matrix metalloproteinases stromelysin 1, and gelatinases A and B (Ki values of 527, 154 and 59 nM, respectively).Ro 32-3555 inhibited interleukin-1α (IL-1α)-induced cartilage collagen degradation in vitro in bovine nasal cartilage explants (IC50=60 nM).Ro 32-3555 was well absorbed in rats when administered orally. Systemic exposure was dose related, with an oral bioavailability of 26% at a dose of 25 mg kg−1.Ro 32-3555 prevented granuloma-induced degradation of bovine nasal cartilage cylinders implanted subcutaneously into rats (ED50=10 mg kg−1, twice daily, p.o.).Ro 32-3555 dosed once daily for 14 days at 50 mg kg−1, p.o., inhibited degradation of articular cartilage in a rat monoarthritis model induced by an intra-articular injection of Propionibacterium acnes.Ro 32-3555 is a potential therapy for the treatment of the chronic destruction of articulating cartilage in both rheumatoid and osteoarthritis.
PMCID: PMC1564706  PMID: 9179398
Cartilage protection agent; collagenase inhibitor; arthritis; cartilage explants; matrix metalloproteinases; pharmacokinetics
6.  Demonstration of carbon-carbon bond cleavage of acetyl coenzyme A by using isotopic exchange catalyzed by the CO dehydrogenase complex from acetate-grown Methanosarcina thermophila. 
Journal of Bacteriology  1991;173(2):929-932.
The purified nickel-containing CO dehydrogenase complex isolated from methanogenic Methanosarcina thermophila grown on acetate is able to catalyze the exchange of [1-14C] acetyl-coenzyme A (CoA) (carbonyl group) with 12CO as well as the exchange of [3'-32P]CoA with acetyl-CoA. Kinetic parameters for the carbonyl exchange have been determined: Km (acetyl-CoA) = 200 microM, Vmax = 15 min-1. CoA is a potent inhibitor of this exchange (Ki = 25 microM) and is formed under the assay conditions because of a slow but detectable acetyl-CoA hydrolase activity of the enzyme. Kinetic parameters for both exchanges are compared with those previously determined for the acetyl-CoA synthase/CO dehydrogenase from the acetogenic Clostridium thermoaceticum. Collectively, these results provide evidence for the postulated role of CO dehydrogenase as the key enzyme for acetyl-CoA degradation in acetotrophic bacteria.
PMCID: PMC207094  PMID: 1987173
7.  In vitro exposure of bovine morulae to Ureaplasma diversum. 
Ureaplasma diversum has been associated with infertility in the cow experimentally and in naturally occurring cases. However, the pathogenic mechanism is undetermined. The purpose of this study was to determine whether ureaplasmas are pathogenic for bovine morulae in vitro. Twenty-one morulae were recovered from three superovulated, mature, Holstein cows six or seven days postestrus. The embryos were divided into three groups (A,B,C) and incubated for 16 hours at 37 degrees C in humidified air with 10% CO2. Group A was incubated in embryo culture medium alone, Group B was incubated in culture medium with sterile ureaplasma broth added and Group C was incubated in culture medium containing 1.7 X 10(6) colony forming units Ureaplasma diversum strain 2312. After incubation, the morulae were examined using an electron microscope. Structures morphologically identical to U. diversum were present on the outer surface of the zonae pellucidae of all the morulae exposed to the organism and none were present on the unexposed control embryos. No other morphological differences were observed in either the ureaplasma-exposed embryos or the two groups of control embryos. Ureaplasma diversum was isolated from three of the five embryos incubated in culture medium with sterile ureaplasma broth added. These three embryos were recovered from one donor cow which cultured positive for U. diversum from the vulva and flush fluid. This finding suggests that the contaminating organisms entered the embryo culture wells either in the embryo collection medium or attached to the embryos.(ABSTRACT TRUNCATED AT 250 WORDS)
PMCID: PMC1255303  PMID: 3607652
8.  Electron microscopy of nickel-containing methanogenic enzymes: methyl reductase and F420-reducing hydrogenase. 
Journal of Bacteriology  1987;169(2):718-727.
Methanogens catalyze the hydrogen-dependent eight-electron reduction of carbon dioxide to methane. Two of the key catalysts in the eight-electron reduction pathway are the nickel-containing enzymes F420-reducing hydrogenase and methyl reductase. In the present study, the structures of these archaebacterial enzymes from Methanobacterium thermoautotrophicum delta H have been determined by electron microscopy. By negative stain techniques, F420 hydrogenase was found to be a ring structure with a diameter of 15.7 nm and an inner channel 4 nm in diameter. Shadow-casting experiments demonstrated that the rings were 8.5 nm deep, indicating a holoenzyme molecular weight of 8.0 X 10(5). Methyl reductase appeared to be an oligomeric complex of dimensions 8.5 by 9 by 11 nm, with a central stain-penetrating region. The morphology and known subunit composition suggest a model in which the subunits are arranged as an eclipsed pair of open trimers. Methyl reductase was also found in the form of larger aggregates and in paracrystalline arrays derived from highly concentrated solutions. The extremely large size of F420 hydrogenase and the methyl reductase supramolecular assemblies may have relevance in vivo in the construction of multiprotein arrays that function in methane biogenesis.
PMCID: PMC211839  PMID: 3804976
9.  Termination of Pregnancy with Cloprostenol and Dexamethasone in Intact or Ovariectomized Cows 
The Canadian Veterinary Journal  1981;22(9):288-290.
Termination of pregnancy in cows was investigated using sham-operated (SH) or ovariectomized (OV) cows treated with either a saline vehicle (V), cloprostenol (PG), dexamethasone (DEX) or dexamethasone and cloprostenol (DEX+PG). Surgery was done at 210 days of pregnancy and treatment was administered 72 hours later.
Days (mean±S.E.) from treatment to termination of pregnancy for the treatment groups were: sham-operated +vehicle (SH+V): 61.5±11.3; ovariectomized+vehicle (OV+V): 53.4±15.7; sham-operated+cloprostenol (SH+PG): 61.8±1.7; ovariectomized+cloprostenol (OV+PG): 54.5±13.1; shamoperated+dexamethasone (SH+DEX): 74.8±4.8; ovariectomized+dexamethasone (OV+DEX): 2.8±0.4; shamoperated+dexamethasone+cloprostenol (SH+DEX+PG) 26.0±23.0; ovariectomized+dexamethasone+cloprostenol (OV+DEX+PG): 7.2±4.9. Pregnancies in the OV+DEX and OV+DEX+PG groups were terminated significantly earlier than in all other groups (P<0.05) except the SH+DEX+PG group. These findings suggest that dexamethasone will terminate pregnancy in cows near seven months of gestation after the ovarian source of progesterone has been removed by either an injection of prostaglandin or by ovariectomy.
PMCID: PMC1789974  PMID: 7343076
11.  Segmental aplasia of the left paramesonephric duct in the cow. 
The Canadian Veterinary Journal  1999;40(12):884-885.
Segmental aplasia of the left uterine horn in a multiparous Holstein cow was diagnosed by palpation and ultrasonography. Treatment with prostaglandin was unsuccessful in eliminating the fluid from the distended uterine horn. Segmental aplasia should be included in the list of differential diagnoses for cows with nonresponsive uterine enlargement.
PMCID: PMC1539888  PMID: 10646066
12.  Measurement of the cytotoxic effects of different strains of Mycoplasma equigenitalium on the equine uterine tube using a calmodulin assay. 
The cytopathic effects induced by five strains of Mycoplasma equigenitalium for cells of equine uterine tube explants were tested by measuring changes in cellular and extracellular concentrations of calmodulin (CaM). Calmodulin concentrations in samples of total homogenate (TH) and total homogenate supernates (THS) of the infected equine uterine tube explants were significantly lower than respective measurements on noninfected controls. In tissue culture medium fractions (TCM) of some infected explants, CaM concentrations were significantly higher than noninfected controls (p > 0.95). The results suggest that M. equigenitalium colonization on ciliated cells of the equine uterine tube can affect the permeability of the cell membrane leading to leakage or release of CaM during cell breakdown. Measurement of CaM concentrations in samples of TH revealed significant differences in the cytotoxic effects induced by different strains of M. equigenitalium on the equine uterine tube (EUT). The data suggests that some strains of M. equigenitalium may have a role in reproductive failure in the mare. In addition comparisons of the means of the concentrations of CaM in samples of TH or THS in EUT explants from four mares in the follicular and four in the luteal phase of the estrous cycle were found to be not significantly different.
PMCID: PMC1263565  PMID: 1477802
13.  Endometrial biopsy in Holstein-Friesian dairy cows. I. Technique, histological criteria and results. 
Endometrial biopsies were taken for histological assessment from 97 cows which calved in a commercial dairy herd between April and August 1984. Sixty-two cows were biopsied at both day 26 and 40 postpartum, 23 cows at only day 26, and 12 at day 40 only. Subjective and quantitative histological criteria were assessed. Ninety-five percent of biopsies were adequate for at least subjective assessment. The distribution of criteria within each horn-day category, as well as combined readings by day and by gravid or nongravid horn were computed and significant differences noted. There was more severe inflammation and more segmented cells at day 26 than 40 postpartum, and in the gravid compared to the nongravid horn. The distribution patterns for the criteria examined provide an overview of histological characteristics in this group of postpartum cows.
PMCID: PMC1263436  PMID: 1884295
14.  Origin of hydrogen in methane produced by Methanobacterium thermoautotrophicum. 
Journal of Bacteriology  1980;141(2):694-698.
The production of deuterated methane by Methanobacterium thermoautotrophicum in H2O-D2O mixtures was examined by high-resolution mass spectrometry. The hydrogen in the methane arose solely from water and not from hydrogen gas. Hydrogen gas served only as an electron source in methanogenesis. A whole-cell product isotope discrimination of 1.5 favoring hydrogen over deuterium was observed in methane production in 81 atom% deuterated water. The distribution of deuterated methane species is described by a simple model of the overall reaction.
PMCID: PMC293677  PMID: 7364716

Results 1-15 (15)