SOME GENETIC AND PHYSIOLOGICAL CHARACTERISTICS OF UREASE-DEFECTIVE STRAINS OF NEUROSPORA CRASSA

1971 ◽  
Vol 13 (2) ◽  
pp. 256-269 ◽  
Author(s):  
Philip Haysman ◽  
H. Branch Howe Jr.
Keyword(s):  
Linkage Group ◽  
Nitrogen Source ◽  
Urease Activity ◽  
Temperature Sensitive ◽  
Sole Nitrogen Source ◽  
Group V ◽  
Group I ◽  
Mutant Strains

Fourteen mutant strains have been isolated which differ from wild type with respect to urease activity: Seven strains lack detectable activity in vivo and in vitro, B1, C21 and allele 601, D1 and allele D3, D2, and W2, and fail to grow with urea as sole nitrogen source; seven have activity in vivo and varying amounts in vitro, A7, E3, E7, R2, S3, and temperature-sensitive strains C5 and K3. Strains D1 and D3 are allelic to Kolmark's ure-1; W2, to Kolmark's ure-2. Strain D2, which is either allelic or closely linked to ure-1, complements none of the strains lacking urease activity nor three of the strains having defective activity, and may be a regulatory mutant. Strains B1, C21 and allele 601 represent two previously unreported loci in linkage group I. Only two of the seven swains having defective urease activity have been mapped, A7 and S3, and have been assigned to linkage group V. These seven strains grow readily on Vogel's medium modified by having urea as sole nitrogen source but not on W-M medium similarly modified; growth is restricted on modified Vogel's medium as well, however, if the initial concentration of nitrogen, as urea, is suitably adjusted to exceed that of phosphorus.

scholarly journals The eutT Gene of Salmonella enterica Encodes an Oxygen-Labile, Metal-Containing ATP:Corrinoid Adenosyltransferase Enzyme

2004 ◽  
Vol 186 (17) ◽  
pp. 5708-5714 ◽  
Author(s):  
Nicole R. Buan ◽  
Sang-Jin Suh ◽  
Jorge C. Escalante-Semerena
Keyword(s):  
Nitrogen Source ◽  
Salmonella Enterica ◽  
Metal Ion ◽  
Complete Loss ◽  
Cell Extracts ◽  
Mutant Strains ◽  
Catalytic Role ◽  
Necessary And Sufficient

ABSTRACT The eutT gene of Salmonella enterica was cloned and overexpressed, and the function of its product was established in vivo and in vitro. The EutT protein has an oxygen-labile, metal-containing ATP:co(I)rrinoid adenosyltransferase activity associated with it. Functional redundancy between EutT and the housekeeping ATP:co(I)rrinoid adenosyltransferase CobA enzyme was demonstrated through phenotypic analyses of mutant strains. Lack of CobA and EutT blocked ethanolamine utilization. EutT was necessary and sufficient for growth of an S. enterica cobA eutT strain on ethanolamine as a carbon and energy or nitrogen source. A eutT+ gene provided in trans corrected the adenosylcobalamin-dependent transcription of a eut-lacZ operon fusion in a cobA strain. Cell extracts enriched for EutT protein contained strong, readily detectable ATP:co(I)rrinoid adenosyltransferase activity. The activity was only detected in extracts maintained under anoxic conditions, with complete loss of activity upon exposure to air or treatment with the Fe2+ ion chelator bathophenanthroline. While the involvement of another metal ion cannot be ruled out, the observed sensitivity to air and bathophenanthroline suggests involvement of Fe2+. We propose that the EutT protein is a unique metal-containing ATP:co(I)rrinoid adenosyltransferase. It is unclear whether the metal ion plays a structural or catalytic role.

scholarly journals Enhancers of conidiation mutants in Aspergillus nidulans.

Genetics ◽  
1994 ◽  
Vol 137 (1) ◽  
pp. 79-85
Author(s):  
D H Gems ◽  
A J Clutterbuck
Keyword(s):  
Linkage Group ◽  
Aspergillus Nidulans ◽  
Double Mutant ◽  
Slight Modification ◽  
Temperature Sensitive ◽  
Permissive Temperature ◽  
Wild Type ◽  
Group V ◽  
Group I

Abstract Mutants at a number of loci, designated sthenyo, have been isolated as enhancers of the oligoconidial mutations at the medA locus. Two loci have been mapped: sthA on linkage group I, and sthB on linkage group V. Two probable alleles have been identified at each locus but two further mutants were unlinked to either sthA or sthB. Neither sthA nor sthB mutants have conspicuous effects on morphology on their own, nor could the sthA1 sthB2 double mutant be distinguished from wild type. Mutants at both loci also interact with the temperature-sensitive brlA42 mutant at the permissive temperature to give a phenotype described as "Abacoid." sthA1 also induces a slight modification of the phenotype of an abaA mutant. We conclude that sthenyo genes act mainly at the phialide stage of conidiation. We also describe the isolation of new medA mutants arising spontaneously as outgrowths on brlA42 colonies.

scholarly journals IN VITRO ANTIOXIDANT POTENTIALS AND HEPATOPROTECTIVE EFFECTS OF Asystasia vogeliana (BENTH) LEAF EXTRACTS AGAINST PARACETAMOL-INDUCED HEPATIC INJURY IN RATS

2020 ◽  
Vol 8 (6) ◽  
pp. 839-848
Author(s):  
Emeka Hillary Ugwuanyi ◽  
◽  
Chukwuneke Udem Samuel ◽  
Ifeanyi Innocent Madubuinyi ◽  
◽  
...  
Keyword(s):  
Antioxidant Activities ◽  
Radical Scavenging ◽  
In Vitro Assays ◽  
Distilled Water ◽  
Leaf Extracts ◽  
Group V ◽  
Reference Drug ◽  
Group I

This study was aimed to investigate the antioxidant potentials of methanol and petroleum ether leaf extracts of Asystasia vogeliana against paracetamol-induced liver injury in rats. For estimation of antioxidant potentials, in vitro radical scavenging assays were carried out using DPPH, FRAP, and ABTS. For in vivo study, twenty-five male Wistar rats weighing 100-120 g were randomized and assigned into 5 groups (I-V, n=5). Further, Paracetamol (PCM) at 2 g/kg was used to induce acute hepatotoxicity orally. Rats in group I received distilled water (10 ml/kg) only. While, the rats of groups II, III, and IV received MLEAV (200 mg/kg), PLEAV (200 mg/kg), and a standard hepatoprotective reference drug silymarin (25 mg/kg) respectively for 5 days before PCM induction. Rats in group V received distilled water for 5 days before PCM induction. Blood and liver samples were collected for hematology, serum biochemistry, and histopathology analyses using standard procedures. In vitro assays revealed that MLEAV showed significant (P < 0.05) increases in antioxidant activity compared with PLEAV. Further, significant (P < 0.05) reductions in the activities of ALT and ALP while a significant (P < 0.05) increases in the activity of antioxidant enzymes (CAT, SOD, and GPx) were reported in the group II and III compared with group V. There were also no observable lesions in their hepatocytes. Results of the study can be concluded that MLEAV elicited more in vitro and in vivo antioxidant activities than PLEAV, thus it protects the liver of rat from PCM-induced hepatotoxicity. Therefore, MLEAV could be used as a hepatoprotective agent for the clinical management of liver damage.

Alcap Delivery Devices and the Sustained Release of Difluoronethylornithine (DFNO) in Vitro and in Vivo Using Male Rats

1987 ◽  
Vol 110 ◽  
Author(s):  
Haned A. Benghuzzi ◽  
Praphulla K. Bajpai
Keyword(s):  
Male Rats ◽  
Control Group ◽  
Sprague Dawley ◽  
Group V ◽  
Group Iii ◽  
Group I ◽  
Body Weights ◽  
Ceramic Implants

AbstractSprague-Dawley albino male rats (25) were divided into five groups consisting of five rats each. Polymer (polylactic acid) impregnated ALCAP capsules filled with 40 mg DFMO were implanted subcutaneously (SC) or intraperitoneally (IP) in Group I and II rats respectively. Rats in Group III were implanted with empty polymer impregnated ALCAP capsules (ALCAP control). Group IV rats were administered orally 3% DFMO in drinking water. Rats in Group V served as controls. Blood samples were collected every week for nine weeks via the tail artery. The concentration of DFNO in the plasma was determined. Data obtained showed that the levels of DFMO in the serum of rats in groups I, I, and IV were 64.71 ±4.08. 219.18 ± 14.48, 16.71 ± 5.21 ug ml−1, respectively at the end of nine weeks. Body weights of the controls and DFMO treated rats were not significantly different (p<0.05). The diarrhea often noted in rats treated orally with DFHO was not observed in rats implanted with ALCAP or ALCAP capsules filled with DFMO. The results of this study suggest that: (1) polymer impregnated ALCAP ceramic implants can be used to deliver DFMO in vivo in a sustained manner for long durations of time, and (2) a ceramic system can be designed to deliver DFNO and drugs such as DFMO in a sustained manner over long durations of time in humans.

Effects of Rest and Exercise on Cardiac Blood Volume Determinations

1997 ◽  
Vol 36 (08) ◽  
pp. 259-264
Author(s):  
N. Topuzović
Keyword(s):  
Radionuclide Ventriculography ◽  
Left Ventricular ◽  
Peak Exercise ◽  
Volume Determination ◽  
Group I ◽  
Lv Volumes ◽  
Group Ii ◽  
Lv Volume

Summary Aim: The purpose of this study was to investigate the changes in blood activity during rest, exercise and recovery, and to assess its influence on left ventricular (LV) volume determination using the count-based method requiring blood sampling. Methods: Forty-four patients underwent rest-stress radionuclide ventriculography; Tc-99m-human serum albumin was used in 13 patients (Group I), red blood cells was labeled using Tc-99m in 17 patients (Group II) in vivo, and in 14 patients (Group III) by modified in vivo/in vitro method. LV volumes were determined by a count-based method using corrected count rate in blood samples obtained during rest, peak exercise and after recovery. Results: In group I at stress, the blood activity decreased by 12.6 ± 5.4%, p <0.05, as compared to the rest level, and increased by 25.1 ± 6.4%, p <0.001, and 12.8 ± 4.5%, p <0.05, above the resting level in group II and III, respectively. This had profound effects on LV volume determinations if only one rest blood aliquot was used: during exercise, the LV volumes significantly decreased by 22.1 ± 9.6%, p <0.05, in group I, whereas in groups II and III it was significantly overestimated by 32.1 ± 10.3%, p <0.001, and 10.7 ± 6.4%, p <0.05, respectively. The changes in blood activity between stress and recovery were not significantly different for any of the groups. Conclusion: The use of only a single blood sample as volume aliquot at rest in rest-stress studies leads to erroneous estimation of cardiac volumes due to significant changes in blood radioactivity during exercise and recovery.

scholarly journals Evaluation of the Safety and Immune Efficacy of Recombinant Human Respiratory Syncytial Virus Strain Long Live Attenuated Vaccine Candidates

2021 ◽  
Author(s):  
Li-Nan Wang ◽  
Xiang-Lei Peng ◽  
Min Xu ◽  
Yuan-Bo Zheng ◽  
Yue-Ying Jiao ◽  
...  
Keyword(s):  
Respiratory Syncytial Virus ◽  
Gene Deletion ◽  
Human Respiratory Syncytial Virus ◽  
Temperature Sensitive ◽  
T7 Promoter ◽  
Vaccine Candidates ◽  
Rsv Infection ◽  
Syncytial Virus

AbstractHuman respiratory syncytial virus (RSV) infection is the leading cause of lower respiratory tract illness (LRTI), and no vaccine against LRTI has proven to be safe and effective in infants. Our study assessed attenuated recombinant RSVs as vaccine candidates to prevent RSV infection in mice. The constructed recombinant plasmids harbored (5′ to 3′) a T7 promoter, hammerhead ribozyme, RSV Long strain antigenomic cDNA with cold-passaged (cp) mutations or cp combined with temperature-sensitive attenuated mutations from the A2 strain (A2cpts) or further combined with SH gene deletion (A2cptsΔSH), HDV ribozyme (δ), and a T7 terminator. These vectors were subsequently co-transfected with four helper plasmids encoding N, P, L, and M2-1 viral proteins into BHK/T7-9 cells, and the recovered viruses were then passaged in Vero cells. The rescued recombinant RSVs (rRSVs) were named rRSV-Long/A2cp, rRSV-Long/A2cpts, and rRSV-Long/A2cptsΔSH, respectively, and stably passaged in vitro, without reversion to wild type (wt) at sites containing introduced mutations or deletion. Although rRSV-Long/A2cpts and rRSV-Long/A2cptsΔSH displayed  temperature-sensitive (ts) phenotype in vitro and in vivo, all rRSVs were significantly attenuated in vivo. Furthermore, BALB/c mice immunized with rRSVs produced Th1-biased immune response, resisted wtRSV infection, and were free from enhanced respiratory disease. We showed that the combination of ΔSH with attenuation (att) mutations of cpts contributed to improving att phenotype, efficacy, and gene stability of rRSV. By successfully introducing att mutations and SH gene deletion into the RSV Long parent and producing three rRSV strains, we have laid an important foundation for the development of RSV live attenuated vaccines.

scholarly journals Group V Phospholipase A2 Mediates Endothelial Dysfunction and Acute Lung Injury Caused by Methicillin-Resistant Staphylococcus aureus

Cells ◽  
2021 ◽  
Vol 10 (7) ◽  
pp. 1731
Author(s):  
Yu Maw Htwe ◽  
Huashan Wang ◽  
Patrick Belvitch ◽  
Lucille Meliton ◽  
Mounica Bandela ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Acute Lung Injury ◽  
Endothelial Dysfunction ◽  
Lung Injury ◽  
Phospholipase A2 ◽  
Methicillin Resistant Staphylococcus Aureus ◽  
Group V ◽  
Methicillin Resistant

Lung endothelial dysfunction is a key feature of acute lung injury (ALI) and clinical acute respiratory distress syndrome (ARDS). Previous studies have identified the lipid-generating enzyme, group V phospholipase A2 (gVPLA2), as a mediator of lung endothelial barrier disruption and inflammation. The current study aimed to determine the role of gVPLA2 in mediating lung endothelial responses to methicillin-resistant Staphylococcus aureus (MRSA, USA300 strain), a major cause of ALI/ARDS. In vitro studies assessed the effects of gVPLA2 inhibition on lung endothelial cell (EC) permeability after exposure to heat-killed (HK) MRSA. In vivo studies assessed the effects of intratracheal live or HK-MRSA on multiple indices of ALI in wild-type (WT) and gVPLA2-deficient (KO) mice. In vitro, HK-MRSA increased gVPLA2 expression and permeability in human lung EC. Inhibition of gVPLA2 with either the PLA2 inhibitor, LY311727, or with a specific monoclonal antibody, attenuated the barrier disruption caused by HK-MRSA. LY311727 also reduced HK-MRSA-induced permeability in mouse lung EC isolated from WT but not gVPLA2-KO mice. In vivo, live MRSA caused significantly less ALI in gVPLA2 KO mice compared to WT, findings confirmed by intravital microscopy assessment in HK-MRSA-treated mice. After targeted delivery of gVPLA2 plasmid to lung endothelium using ACE antibody-conjugated liposomes, MRSA-induced ALI was significantly increased in gVPLA2-KO mice, indicating that lung endothelial expression of gVPLA2 is critical in vivo. In summary, these results demonstrate an important role for gVPLA2 in mediating MRSA-induced lung EC permeability and ALI. Thus, gVPLA2 may represent a novel therapeutic target in ALI/ARDS caused by bacterial infection.

scholarly journals Important Role for Phylogenetically Invariant PP2Acα Active Site and C-Terminal Residues Revealed by Mutational Analysis in Saccharomyces cerevisiae

Genetics ◽  
2000 ◽  
Vol 156 (1) ◽  
pp. 21-29 ◽  
Author(s):  
David R H Evans ◽  
Brian A Hemmings
Keyword(s):  
Protein Interactions ◽  
Active Site ◽  
Protein Function ◽  
Mutational Analysis ◽  
Yeast Cells ◽  
Temperature Sensitive ◽  
Active Site Residues ◽  
Catalytic Function

Abstract PP2A is a central regulator of eukaryotic signal transduction. The human catalytic subunit PP2Acα functionally replaces the endogenous yeast enzyme, Pph22p, indicating a conservation of function in vivo. Therefore, yeast cells were employed to explore the role of invariant PP2Ac residues. The PP2Acα Y127N substitution abolished essential PP2Ac function in vivo and impaired catalysis severely in vitro, consistent with the prediction from structural studies that Tyr-127 mediates substrate binding and its side chain interacts with the key active site residues His-118 and Asp-88. The V159E substitution similarly impaired PP2Acα catalysis profoundly and may cause global disruption of the active site. Two conditional mutations in the yeast Pph22p protein, F232S and P240H, were found to cause temperature-sensitive impairment of PP2Ac catalytic function in vitro. Thus, the mitotic and cell lysis defects conferred by these mutations result from a loss of PP2Ac enzyme activity. Substitution of the PP2Acα C-terminal Tyr-307 residue by phenylalanine impaired protein function, whereas the Y307D and T304D substitutions abolished essential function in vivo. Nevertheless, Y307D did not reduce PP2Acα catalytic activity significantly in vitro, consistent with an important role for the C terminus in mediating essential protein-protein interactions. Our results identify key residues important for PP2Ac function and characterize new reagents for the study of PP2A in vivo.

scholarly journals Effects of bisphosphonate and calcium carbonate on normal aortic valve and cholecalciferol induced in vivo rabbit aortic stenosis model

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
S Okutucu ◽  
C Sabanoglu ◽  
A Saglam Ayhan ◽  
E Tulumen ◽  
H Aksoy ◽  
...  
Keyword(s):  
Calcium Carbonate ◽  
Aortic Valve ◽  
Aortic Stenosis ◽  
Calcium Supplementation ◽  
Group Iv ◽  
Histopathologic Examination ◽  
Group V ◽  
Group I ◽  
Alendronate Treatment

Abstract Background Calcific aortic valve disease (CAVD) is the most common valvular heart disease. Bisphosphonates are stable analogs of pyrophosphates and commonly prescribed in the treatment of osteoporosis. The effects of bisphosphonate treatment on CAVD are not clearly known and there are inconsistent results. Similarly, the effect of calcium supplementation on CAVD remains controversial. Purpose The aim of this study was to assess the effects of bisphosphonate therapy on the normal aortic valve and vitamin D induced in vivo rabbit aortic stenosis (AS) model. Methods The impact of calcium supplementation on the rabbit AS model was also evaluated. A total of 30 New Zealand white rabbits were divided into five equal groups: no treatment (Group I); 25,000 IU/day vitamin D3 (cholecalciferol) (Group II, rabbit AS model); 25,000 IU/day cholecalciferol plus 2500 mg/day calcium carbonate (Group III); 20 μg/kg/week intravenous alendronate (Group IV) and 25,000 IU/day cholecalciferol plus 2500 mg/day calcium carbonate plus 20μg/kg/week alendronate (Group V). Echocardiography was performed at baseline and after 12 weeks of treatment. The left ventricular mass index (LVMI), aortic valve area (AVA), transvalvular velocities and gradients were recorded. Radiologic and histopathologic examination was performed at the end of the 12th week. Control animals displayed no abnormalities of the aortic valve. Results There was no echocardiographic change in Group IV. In Groups II, III and V, there was a significant decrease in AVA and increases in transvalvular velocities and gradients. However, these stenotic changes were significantly prominent in Group V (p=0.001 for all, via repeated measures ANOVA). Moreover, LVMI was only increased in Group V (p&lt;0.05). Calcification of aortic valvar complex was detected in 14 (46.7%) cases by radiologic imaging and 10 (33.3%) cases by histopathologic examination. Most frequent calcification was found in Group V (5 for each method, 83.3%). Agatston, volume and equivalent mass scores of calcific foci in Group V were significantly higher than other groups (p&lt;0.05 for all). There was no significant difference between groups regarding with presence of osteoclasts in calcific foci. Conclusion Calcium supplementation has no effect on the in vivo rabbit AS model. Alendronate treatment aggravates the stenosis and increases the calcification in the rabbit AS model. Alendronate treatment has no effect on the normal valve in which there was no osteogenesis and osteoclastogenesis. Based on these findings, in patients with CAVD, alendronate treatment should be given with regular echocardiographic follow-up or may not be preferred. Central figure Funding Acknowledgement Type of funding source: None

scholarly journals C-Terminal Deletions Can Suppress Temperature-Sensitive Mutations and Change Dominance in the Phage Mu Repressor

Genetics ◽  
1996 ◽  
Vol 142 (3) ◽  
pp. 661-672 ◽  
Author(s):  
Jodi L Vogel ◽  
Vincent Geuskens ◽  
Lucie Desmet ◽  
N Patrick Higgins ◽  
Ariane Toussaint
Keyword(s):  
Binding Activity ◽  
Temperature Sensitive ◽  
Dna Binding Activity ◽  
Heat Stable ◽  
C Terminus ◽  
Acid Domain ◽  
Mutant Forms ◽  
Amino Acid Domain

Abstract Mutations in an N-terminal 70-amino acid domain of bacteriophage Mu's repressor cause temperature-sensitive DNA-binding activity. Surprisingly, amber mutations can conditionally correct the heat-sensitive defect in three mutant forms of the repressor gene, cts25 (D43-G), cts62 (R47-Q and cts71 (M28-I), and in the appropriate bacterial host produce a heat-stable Sts phenotype (for survival of temperature shifts). Sts repressor mutants are heat sensitive when in supE or supF hosts and heat resistant when in Sup° hosts. Mutants with an Sts phenotype have amber mutations at one of three codons, Q179, Q187, or Q190. The Sts phenotype relates to the repressor size: in Sup° hosts sts repressors are shorter by seven, 10, or 18 amino acids compared to repressors in supE or supF hosts. The truncated form of the sts62-1 repressor, which lacks 18 residues (Q179–V196), binds Mu operator DNA more stably at 42° in vitro compared to its full-length counterpart (cts62 repressor). In addition to influencing temperature sensitivity, the C-terminus appears to control the susceptibility to in vivo Clp proteolysis by influencing the multimeric structure of repressor.

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