Biomedical subjects

V Aguilar

Publications and source records attributed to V Aguilar.

10 recordsLinked to original sources

PubMed1990

The tryptophan synthase bienzyme complex transfers indole between the alpha- and beta-sites via a 25-30 A long tunnel.

The bacterial tryptophan synthase bienzyme complexes (with subunit composition alpha 2 beta 2) catalyze the last two steps in the biosynthesis of L-tryptophan. For L-tryptophan synthesis, indole, the common metabolite, must be transferred by some mechanism from the alpha-catalytic site to the beta-catalytic site. The X-ray structure of the Salmonella typhimurium tryptophan synthase shows the catalytic sites of each alpha-beta subunit pair are connected by a 25-30 A long tunnel [Hyde, C. C., Ahmed, S. A., Padlan, E. A., Miles, E. W., & Davies, D. R. (1988) J. Biol. Chem. 263, 17857-17871]. Since the S. typhimurium and Escherichia coli enzymes have nearly identical sequences, the E. coli enzyme must have a similar tunnel. Herein, rapid kinetic studies in combination with chemical probes that signal the bond formation step between indole (or nucleophilic indole analogues) and the alpha-aminoacrylate Schiff base intermediate, E(A-A), bound to the beta-site are used to investigate tunnel function in the E. coli enzyme. If the tunnel is the physical conduit for the transfer of indole from the alpha-site to the beta-site, then ligands that block the tunnel should also inhibit the rate at which indole and indole analogues from external solution react with E(A-A). We have found that when D,L-alpha-glycerol 3-phosphate (GP) is bound to the alpha-site, the rate of reaction of indole and nucleophilic indole analogues with E(A-A) is strongly inhibited. These compounds appear to gain access to the beta-site via the alpha-site and the tunnel, and this access is blocked by the binding of GP to the alpha-site. However, when small nucleophiles such as hydroxylamine, hydrazine, or N-methylhydroxylamine are substituted for indole, the rate of quinonoid formation is only slightly affected by the binding of GP. Furthermore, the reactions of L-serine and L-tryptophan with alpha 2 beta 2 show only small rate effects due to the binding of GP. From these experiments, we draw the following conclusions: (1) L-Serine and L-tryptophan gain access to the beta-site of alpha 2 beta 2 directly from solution. (2) The small effects of GP on the rates of the L-serine and L-tryptophan reactions are due to GP-mediated allosteric interactions between the alpha- and beta-sites.(ABSTRACT TRUNCATED AT 400 WORDS)

Allosteric Regulation↗

PubMed1990

Hemispheric local/global processing revisited.

Hemispheric differences for local and global processing were assessed with square and rectangular stimulus shapes which contained either squares or rectangles within them. Experiment 1 manipulated the overall size and area of the stimulus items, while experiment 2 kept stimulus area constant and manipulated the number of shapes inside the stimulus. In different conditions subjects judged whether the inside shapes (local processing) or whether the outside shapes (global processing) were squares or rectangles. Reaction time measures revealed that hemispheric differences were unrelated to the local or global processing requirements as well as the spatial frequency of the stimulus texture. A left hemisphere advantage generally was obtained across stimulus and processing conditions, with the pattern of task effects dependent on the specific stimuli employed. The results suggest that the overall stimulus shape rather than processing mode or specific spatial frequency is a primary determinant of hemispheric differences for visual information.

Adult↗

PubMed1990

[Cytometric profiles of DNA content in cervical intraepithelial neoplasia (CIN)].

The cervical cytologic screening is responsible for the overall decline in the incidence of cervical cancer. Despite this apparent success, the cytologic assessment by light microscopy of cervical smear has recently been shown to have deficiencies particularly in cervical intraepithelial neoplasia (CIN). Now, this pathology of the uterine cervix is increasingly becoming recognized as a major health problem in young women because of its high frequency and its association with the development of cervical carcinoma. Flow cytometry (FCM) is widely used as a rapid method to analyse DNA content of a great number of cells on a cell by cell basis. The authors have applied FCM to CIN in order to reinforce the cytologic screening of cervical smears. Three abnormal DNA patterns have been observed: increased and heterogeneous DNA pattern; increased S and G2+M phases; abnormal DNA content. The statistical calculation has emphasized the significant relation linking these DNA patterns with the cytologic diagnosis and the severity of CIN. Therefore, FCM is a powerful complementary tool in cervical pathology.

Carcinoma in Situ↗

PubMed1990

[Qualitative approach of DNA content analysis by flow cytometry].

One step DNA staining procedures are very easy to use. Reduced staining time and small amount of needed components explain the world wide use of these protocols. The heterogeneity of the results obtained with these technics during a study of different mice organs has let the authors to think that there was a close relationship between this heterogeneity and DNA staining technics. The authors have observed the same discordances with some human tissues (parathyroid, prostatic adenoma...). They disappear when the same cells are stained with a procedure that associates to propidium iodide and RNAse a pretreatment of nuclei by trypsin and spermin. These molecules are DNA stabilizing components. So one can develop the hypothesis that the abnormalities observed with the one step procedures are due to a decondensated state of DNA that allows the fixation of a higher amount of stain. The main point will be to find a direct relationship with different physiological or pathological states. Now, it is necessary to sensibilise users against the risk of false abnormal DNA content detection through the one step procedures.

Animals↗

PubMed1988

[Inter-organ differences of the cytometric DNA content in mice: relation of the staining method].

With one step DNA staining methods including cell membrane lysis and RNase treatment, we regularly observed a higher fluorescence emission in liver nuclei compared to bone marrow nuclei in C57BL/6 mice. Therefore this study was conducted in order to emphasize such a phenomenon in other organs and to assess if higher fluorescence emission was related to higher DNA content or staining procedure failure. Liver, bone marrow and testis were removed from Swiss, BDF and C57BL/6 mice. The following samples were prepared: 1) liver cells with TRBC (TRBC = Trout Red Blood Cells = internal standards), 2) bone marrow cells with TRBC, 3) testis cells with TRBC and 4) mixtures of liver, bone marrow and testis cells. The staining procedures were: A) one step pH 10 procedure described by Vindelov (Virchows Arch. B. Cell Path., 1977, 24, 227-242), B) same procedure with twice RNase concentration, C) first method with twice NP 40 concentration and D) three steps procedure including Trypsin and Spermine treatment (Vindelov et al., Cytometry, 1983, 3, 323-327). In protocols A, B and C, "Diploid cells/TRBC" ratio differed significantly between liver, bone marrow and testis nuclei. Moreover, 3 distinct populations of diploid cells were present in samples 4. In protocol D, "Diploid cells/TRBC" ratio were identical between liver, bone marrow and testis nuclei. In samples 4, only 1 population of diploid cells has been observed. This study shows that DNA stabilization by polyamine and protein degradation by protease could act on Propidium Iodide fixation and/or fluorescence emission, with significant differences according to the origin of the cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

PubMed1988