ABSTRACT
PC2 prohormone convertases are enzymes involved in the proteolytic maturation of neuropeptide precursors. In the present work, a cDNA encoding a PC2-like enzyme (OrlPC2) was cloned from crayfish eyestalk ganglia (medulla terminalis) containing the X-organ, a major neuroendocrine center. The predicted 634 amino acid preproprotein exhibits highest sequence identity, especially in the catalytic domain, with PC2s from arthropods and nematodes, and less with mollusc and vertebrate enzymes. It was demonstrated by in situ hybridization on crayfish medulla terminalis sections that OrlPC2 is expressed in a large number of neuron perikarya, including those producing the well known crustacean hyperglycemic hormone.
Subject(s)
Astacoidea/genetics , Subtilisins/genetics , Amino Acid Sequence , Animals , Base Sequence , Cloning, Molecular , DNA Primers , DNA, Complementary/biosynthesis , DNA, Complementary/chemistry , In Situ Hybridization , Molecular Sequence Data , Proprotein Convertase 2 , Sequence Alignment , Subtilisins/biosynthesis , Subtilisins/chemistryABSTRACT
Biochemical studies on ecdysteroid metabolism in arthropods suggest that aldoketoreductase enzymes (AKRs) may be involved in this pathway, but very few molecular data are available on these oxidoreductases in invertebrates. Looking for such enzymes in the crayfish Orconectes limosus, we have used a PCR strategy with primers deduced from a recent insect 3beta-reductase sequence, and from mammalian 5beta-reductase sequences. A full-length cDNA, corresponding to a putative AKR, was isolated from crayfish antennal gland. This cDNA contains an open-reading frame of 1008 bp, encoding a predicted protein of 336 amino acids. Northern blots indicated a restricted expression of the transcript in the antennal glands, quite constant during the molting cycle, and in situ hybridization demonstrated a strong expression of the transcript in the labyrinth. This is to date the first member of the AKRs superfamily characterized in a crustacean species, and the putative function of the corresponding enzyme is discussed.
Subject(s)
Alcohol Oxidoreductases/genetics , Astacoidea/genetics , Gene Expression , Aldehyde Reductase , Aldo-Keto Reductases , Amino Acid Sequence , Animals , Astacoidea/anatomy & histology , Astacoidea/enzymology , Base Sequence , Blotting, Northern , DNA, Complementary , In Situ Hybridization , Molecular Sequence Data , Open Reading Frames , Reverse Transcriptase Polymerase Chain Reaction , Sequence Homology, Amino AcidABSTRACT
1. This investigation was undertaken for the purpose of determining the structure of the phosphatidylcholines of lung surfactant system present at birth in normal full term newborn infants. 2. The procedure, using tracheal aspirates as lung secretions, combines a cold-acetone precipitation and a two-dimensional thin-layer chromatography of the lipid extract. 3. Different species of phosphatidylcholines were isolated and found to account together for over 60% of the total phospholipids in tracheal aspirates. Analysis of the fatty acids esterifying the alpha- and beta-carbon of these different phosphatidylcholines showed palmitic acid as the major component with little myristic acid. 4. This fatty acid analysis revealed furthermore that the major phosphatidylcholine fraction was almost exclusively alpha, beta-dipalmitoylphosphatidylcholine. 5. This study shows that the procedure described provides a useful and simple method for the extraction, isolation and characterisation of the functional components of lung surfactant in living human newborns.
