CGRP 27-37 analogues with high affinity to the CGRP1 receptor show antagonistic properties in a rat blood flow assay

CGRP Y0-28-37 is known as a selective CGRP1 receptor antagonist. We succeeded in optimising the CGRP1 receptor affinity of this fragment by multiple amino acid replacement. The analogues [p34, F35]CGRP 27-37 and [D31, p34, F35]CGRP 27-37 exhibit a 100-fold increased affinity compared to the unmodified segment. Receptor binding studies were performed with human neuroblastoma cells SK-N-MC, which selectively express the hCGRP1 receptor. Blood flow, which is increased by exogenous CGRP, was measured in the right femoral artery. Preincubation of the rats with [p34, F35]CGRP 27-37 and [D31, p34, F35]CGRP 27-37 led to a significant decrease in CGRP induced increase in vascular conductance indicating the antagonistic properties of these compounds. Interestingly, an exchange of the amino acid Asn31 to Asp31 in [p34, F35]CGRP 27-37 shortened the period of the antagonistic effect significantly, suggestive of a different rate of metabolism for the two ligands. Secondary structure investigations obtained by circular dichroism measurements revealed that an increase in ordered structure correlates with high binding affinity.     

Changes in growth,proteins and free amino acids of developing seed and pod of fenugreek

The growth and development under field conditions of the pods and seeds of two cvs of Trigonella foenum graecum are described. Samples were harvested at different stages of ripeness for the determination of dry matter, protein and free amino acid content. During maturation, reserves of solutes are established in the pod wall before the seeds begin their exponential phase of growth. Later, these reserves disappear, providing about 20% of the seed's requirements for nitrogen. SDS-electrophoresis was used to follow the formation of proteins and it was shown that the synthesis of storage proteins takes place prior to dehydration of the seed. Production soluble nitrogenous compounds precedes protein accumulation. Free amino acids follow the same pattern. 4-Hydroxyisoleucine represents nearly 80% of free amino acid of dry seeds. The concentration does not decrease in the later stages of maturation of the seed but this unusual amino acid is absent in the storage proteins of the seeds.     

Cell-Type Specific Four-Component Hydrogel

In the field of regenerative medicine we aim to develop implant matrices for specific tissue needs. By combining two per se, cell-permissive gel systems with enzymatic crosslinkers (gelatin/transglutaminase and fibrinogen/thrombin) to generate a blend (technical term: quattroGel), an unexpected cell-selectivity evolved. QuattroGels were porous and formed cavities in the cell diameter range, possessed gelation kinetics in the minute range, viscoelastic properties and a mechanical strength appropriate for general cell adhesion, and restricted diffusion. Cell proliferation of endothelial cells, chondrocytes and fibroblasts was essentially unaffected. In contrast, on quattroGels neither endothelial cells formed vascular tubes nor did primary neurons extend neurites in significant amounts. Only chondrocytes differentiated properly as judged by collagen isoform expression. The biophysical quattroGel characteristics appeared to leave distinct cell processes such as mitosis unaffected and favored differentiation of sessile cells, but hampered differentiation of migratory cells. This cell-type selectivity is of interest e.g. during articular cartilage or invertebral disc repair, where pathological innervation and angiogenesis represent adverse events in tissue engineering.     

Development of an in vitro bacteriophage N4 DNA replication system

An in vitro DNA replication system from bacteriophage N4-infected Escherichia coli has been developed. It requires MgCl2, all four deoxyribonucleoside triphosphates, and exogenously added N4 phage DNA; other DNAs are used inefficiently or not at all. Ribonucleoside triphosphates are not required, although they stimulate DNA synthesis. In vitro replication starts at the ends of the N4 genome and moves progressively inward. Initiation occurs through hairpin priming at the 3' ends of the genome, but shows a strong preference for the right end. Three N4 gene products (dnp, dbp, and exo) required in vivo for N4 DNA synthesis are absolutely required in the in vitro system. These findings are discussed with respect to the mode of N4 DNA replication.