RESUMO
Infrared spectra of phenyl acridine-9-carboxylates and their 10-methylated cationic derivatives were recorded and discussed. Experimental data were compared with theoretically predicted transitions at the DFT level of theory (using the B3LYP functional and 6-31G** basis set) for optimized geometries of molecules. Substitution influences the values of the wavenumbers of characteristic stretching and bending modes, i.e. those corresponding to ester groups and fragments of molecules containing a heterocyclic nitrogen atom. The experimentally determined transitions of selected groups of atoms correlate well with the theoretically predicted values. Interdependences among some theoretically derived physicochemical features of the compounds and IR frequencies of selected bands are also discussed.
Assuntos
Acridinas/química , Modelos Químicos , Vibração , Cátions , Conformação Molecular , Espectroscopia de Infravermelho com Transformada de FourierRESUMO
The sarco(endo)plasmic reticulum Ca2+ ATPase (SERCA) type 1 and 2 genes are alternatively spliced at their 3' end. We hypothesized that similar mechanism may occur for SERCA 3. Two spliced variants were identified by RNase protection analysis. We then isolated and sequenced the 3' end portion of the mouse SERCA 3 gene, and confirmed the presence of an alternative mRNA transcript by sequencing a cDNA fragment obtained by RT-PCR. Tissue distribution of the alternatively spliced mRNAs was studied by RT-PCR: SERCA 3b was the only isoform expressed in endothelial cells from aorta and heart and also was the major isoform in lung and kidney whereas SERCA 3a and 3b were coexpressed in trachea, intestine, thymus, spleen, and fetal liver.
Assuntos
Processamento Alternativo , ATPases Transportadoras de Cálcio/genética , Isoenzimas/genética , RNA Mensageiro/metabolismo , Retículo Sarcoplasmático/enzimologia , Sequência de Aminoácidos , Animais , Sequência de Bases , ATPases Transportadoras de Cálcio/metabolismo , Expressão Gênica , Isoenzimas/metabolismo , Camundongos , Dados de Sequência Molecular , Especificidade de Órgãos , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , RatosRESUMO
Human platelets express several sarco/endoplasmic reticulum Ca2+ ATPase (SERCA) isoenzymes: SERCA2b of 100 kDa apparent molecular mass and two distinct enzymes of 97 kDa, one of them identified as being the SERCA3a isoform. The molecular identity of the third enzyme specifically recognized by the PL/IM430 monoclonal antibody has remained elusive. First, the study of the 3'-end part of platelet SERCA3 mRNA, by means of RT-PCR amplification using sets of primers covering the N-3 to N (ultimate) exons of the human SERCA3 sequence, revealed the presence of two distinct mRNA sequences, SERCA3a and a longer variant. Second, this additional sequence was identified as SERCA3b and found to refer to the insertion of a new exon of 73 bp, located at bp 349 from the beginning of the intronic sequence, linking the penultimate (N-1) exon to the last exon (N) of the human SERCA3 gene. Third, a relationship between the expression of this SERCA3b mRNA and the PL/ IM430 recognizable SERCA protein was observed. SERCA3b mRNA was found to be absent in epithelial HeLa cells not recognized by the PL/IM430 antibody and the expression of this SERCA3b RNA species correlated with that of the SERCA protein recognized by PL/IM430 which was down-modulated in the platelet precursor megakaryocytic CHRF 288-11 cell line as well as upon in vitro lymphocyte activation. Taken together, these results strongly support the notion of the presence of the SERCA3b protein in human cells by showing SERCA3b mRNA in platelets and the fact that the protein corresponding to this mRNA species is very likely the 97 kDa protein recognized by the PL/IM430 antibody.
Assuntos
Especificidade de Anticorpos , Plaquetas/enzimologia , ATPases Transportadoras de Cálcio/imunologia , ATPases Transportadoras de Cálcio/metabolismo , Retículo Endoplasmático/enzimologia , Isoenzimas/metabolismo , Sequência de Aminoácidos , Anticorpos Monoclonais/imunologia , ATPases Transportadoras de Cálcio/genética , Éxons , Células HeLa , Humanos , Isoenzimas/genética , Isoenzimas/imunologia , Dados de Sequência Molecular , RNA Mensageiro/metabolismo , Homologia de Sequência de AminoácidosRESUMO
The far-ultraviolet circular dichroism and fluorescence emission intensities of the myosin rod were studied at pH 2-11, in the absence and presence of guanidine hydrochloride. The protein kept its helicity in this pH range. Its stability in the denaturant was higher at acidic pH than at pH 7. This may be due to favorable interactions involving protonated glutamic acid residues at the interface of the two alpha-helical chains of the molecule. At alkaline pH, the fluorescence of the myosin rod was quenched, and the tryptophan region of the protein became less stable in the presence of guanidine hydrochloride, due to ionization of tyrosine residues or other amino acids close to tryptophans in the double-helix arrangement.
