Synthesis and modification of the Amyloid peptide sequence 37-42 of Aβ42 (AβPP): Efficient synthesis of N-methylated peptides, expanding the toolsfor peptide research

[EN] We report the synthesis and characterization of N-alkyl modified peptides by efficient coupling of N-methyl amino acids in solution phase. As a model peptide, the segment 37-42 (GGVVIA) of the Aβ-42 amyloid peptide derived from the amyloid precursor protein (Aβ-PP) was chosen. This peptide and its derivatives with N-methyl groups on Val40 and Ile41 residues were synthesized and character-ized. Because the synthesis was performed in solution-phase, the pro-cedure can be easily scaled up for the production of larger amounts of the peptides described in this work or any linear N-methyl peptide with potential therapeutic application.[ES] Se describe aquí la síntesis y caracterización de péptidos N-alquil modificados por acoplamiento eficiente en solución de N-Me-til aminoácidos. Se eligió como péptido modelo el segmento de 37-42 (GGVVIA) del péptido amiloide Aβ-42 derivado de la proteína pre-cursora de amiloide (Aβ-PP). Este péptido y sus derivados con grupos N-metilo en los residuos Val40 y Ile41 fueron sintetizados y caracteri-zados. Dado que ésta síntesis se realizó en solución, el procedimiento puede escalarse fácilmente y producir cantidades suficientes de los péptidos descritos, pudiendo aplicar ésta técnica en otros péptidos N-metilados lineales con posible potencial aplicación terapéutica.We are grateful to CONACYT for financial support (Project CB2010/151875); for scholarship to M.E.R-V. A. B. thanks the Plan Nacional de Investigación, MINECO, Spain and FSE funds for generous support through grant SAF2013-48399-R and Laboratorio Nacional de Estructura de Macromoléculas (LANEM, CONACyT 251613).Peer Reviewe

A central regulatory system largely controls transcriptional activation and repression responses to phosphate starvation in Arabidopsis.

Plants respond to different stresses by inducing or repressing transcription of partially overlapping sets of genes. In Arabidopsis, the PHR1 transcription factor (TF) has an important role in the control of phosphate (Pi) starvation stress responses. Using transcriptomic analysis of Pi starvation in phr1, and phr1 phr1-like (phl1) mutants and in wild type plants, we show that PHR1 in conjunction with PHL1 controls most transcriptional activation and repression responses to phosphate starvation, regardless of the Pi starvation specificity of these responses. Induced genes are enriched in PHR1 binding sequences (P1BS) in their promoters, whereas repressed genes do not show such enrichment, suggesting that PHR1(-like) control of transcriptional repression responses is indirect. In agreement with this, transcriptomic analysis of a transgenic plant expressing PHR1 fused to the hormone ligand domain of the glucocorticoid receptor showed that PHR1 direct targets (i.e., displaying altered expression after GR:PHR1 activation by dexamethasone in the presence of cycloheximide) corresponded largely to Pi starvation-induced genes that are highly enriched in P1BS. A minimal promoter containing a multimerised P1BS recapitulates Pi starvation-specific responsiveness. Likewise, mutation of P1BS in the promoter of two Pi starvation-responsive genes impaired their responsiveness to Pi starvation, but not to other stress types. Phylogenetic footprinting confirmed the importance of P1BS and PHR1 in Pi starvation responsiveness and indicated that P1BS acts in concert with other cis motifs. All together, our data show that PHR1 and PHL1 are partially redundant TF acting as central integrators of Pi starvation responses, both specific and generic. In addition, they indicate that transcriptional repression responses are an integral part of adaptive responses to stress

Three Isoforms of Isoamylase Contribute Different Catalytic Properties for the Debranching of Potato Glucans

Isoamylases are debranching enzymes that hydrolyze α-1,6 linkages in α-1,4/α-1,6–linked glucan polymers. In plants, they have been shown to be required for the normal synthesis of amylopectin, although the precise manner in which they influence starch synthesis is still debated. cDNA clones encoding three distinct isoamylase isoforms (Stisa1, Stisa2, and Stisa3) have been identified from potato. The expression patterns of the genes are consistent with the possibility that they all play roles in starch synthesis. Analysis of the predicted sequences of the proteins suggested that only Stisa1 and Stisa3 are likely to have hydrolytic activity and that there probably are differences in substrate specificity between these two isoforms. This was confirmed by the expression of each isoamylase in Escherichia coli and characterization of its activity. Partial purification of isoamylase activity from potato tubers showed that Stisa1 and Stisa2 are associated as a multimeric enzyme but that Stisa3 is not associated with this enzyme complex. Our data suggest that Stisa1 and Stisa2 act together to debranch soluble glucan during starch synthesis. The catalytic specificity of Stisa3 is distinct from that of the multimeric enzyme, indicating that it may play a different role in starch metabolism

Three isoforms of isoamylase contribute different catalytic properties for the debranching of potato glucans

Isoamylases are debranching enzymes that hydrolyze � -1,6 linkages in � -1,4/ � -1,6–linked glucan polymers. In plants, they have been shown to be required for the normal synthesis of amylopectin, although the precise manner in which they influence starch synthesis is still debated. cDNA clones encoding three distinct isoamylase isoforms (Stisa1, Stisa2, and Stisa3) have been identified from potato. The expression patterns of the genes are consistent with the possibility that they all play roles in starch synthesis. Analysis of the predicted sequences of the proteins suggested that only Stisa1 and Stisa3 are likely to have hydrolytic activity and that there probably are differences in substrate specificity between these two isoforms. This was confirmed by the expression of each isoamylase in Escherichia coli and characterization of its activity. Partial purification of isoamylase activity from potato tubers showed that Stisa1 and Stisa2 are associated as a multimeric enzyme but that Stisa3 is not associated with this enzyme complex. Our data suggest that Stisa1 and Stisa2 act together to debranch soluble glucan during starch synthesis. The catalytic specificity of Stisa3 is distinct from that of the multimeric enzyme, indicating that it may play a different role in starch metabolism