Additionally, these peptides partially competed with formyl-Met-Leu-Phe binding to HL-60 cells differentiated into neutrophilic/granulocytic cells, presenting the possibility that the receptor for MCT-2 and hMCT-2 is one of the formyl peptide receptors. Moreover, MCT-2 and hMCT-2 also stimulated beta-hexosaminidase secretion in human neutrophils isolated from peripheral blood in a concentration-dependent manner.
N TERMINAL SEQUENCE ANALYSIS FREE
The induction of beta-hexosaminidase release, chemotaxis, and the increase in the intracellular free Ca(2+) concentration by hMCT-2 were completely suppressed by pertussis toxin, indicating the involvement of G(i)- or G(o)-type G proteins in the signaling pathways. Synthetic MCT-2 and its human homolog hMCT-2 induced beta-hexosaminidase release in and chemotaxis of HL-60 cells differentiated into neutrophilic/granulocytic cells. Structural analyses indicated that the primary structure of this peptide is formyl-Met-Thr-Asn-Ile-Arg-Lys-Ser-His-Pro-Leu-Met-Lys-Ile-Ile-Asn, which is identical to that of the N-terminal pentadecapeptide of porcine mitochondrial cytochrome b we therefore named the newly isolated peptide "mitocryptide-2" (MCT-2), since we have recently purified and identified mitocryptide-1, a different class of a neutrophil-activating peptide. Here, we isolated and identified from porcine hearts a neutrophil-activating peptide. The actions of neutrophils in damaged tissues are still not well understood, particularly in the early stage of inflammation, and as-yet-unknown neutrophil-activating substances are proposed to induce their acute transmigration and activation. Neutrophils are a class of leukocytes involved in innate immunity by monitoring and scavenging invading microorganisms and toxic substances. Finally, discussion of optimization of sequencer performance as well as possible solutions to frequently encountered problems is included. A discussion of data interpretation is therefore provided. The amount of data obtained from a single sequencer run is substantial, and careful interpretation of this data by an experienced scientist familiar with the current operation performance of the instrument used for this analysis is critically important. Methods are provided for optimizing separation of PTH amino acid derivatives on Perkin-Elmer instruments and for increasing the proportion of sample injected onto the PTH analyzer on older Perkin-Elmer instruments by installing a modified sample loop. Sequence analysis of protein or peptide samples in solution or bound to PVDF membranes using a Hewlett-Packard Model G1005A sequencer is also described. This unit describes the sequence analysis of protein or peptide samples in solution or bound to PVDF membranes using a Perkin-Elmer Procise Sequencer. Amino-terminal (N-terminal) sequence analysis is used to identify the order of amino acids of proteins or peptides, starting at their N-terminal end.
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