By J. R. Vane (auth.), C. N. Serhan, H. D. Perez (eds.)
Over the previous couple of years, we've witnessed large growth within the box of eicosanoids and their healing purposes. Receptor an tagonists for leukotrienes were confirmed as anti-inflammatories and are out there as a therapy for bronchial asthma. Receptor agonists for professional stacyclin are being verified for the remedy of peripheral vascular dis ease, and selective inhibitors of cyclooxygenase sort II have been simply ap proved for the therapy of rheumatoid arthritis. some of these advancements are the fruits of decades and man-hours of cautious learn. the sector has now entered an upswing that may bring about novel thera peutic functions in the subsequent 10 years. New molecules and me diators were pointed out, new enzymes and pathways elucidated and new healing ways have emerged. the idea that of ei cosanoids as "pro-inflammatory" molecules is being challenged, and their function as regulators is more and more famous. in truth, a few of these molecules might be very important endogenous anti inflammatory agents.
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Additional info for Advances in Eicosanoid Research
P. ). Oligonucleotide primers were synthesized by Genosys (Woodlands, Texas). Radiolabeled phospholipids, chromatography resins, restriction enzymes and DNA-mod ifying enzymes were from Amersham Pharmacia Biotech (Piscataway, New Jersey) . All other chemicals used were of at least analytical grade and were obtained from either Fisher or Sigma. 2 Cloning The pCH 10 vector containing the gene for human group- V PLA z was obtained from Jay A. Tischfield of Indiana University. The group-V gene was released by digestion with NheI and Xhol.
To test for activ ation by anionic surfaces, 5 mole percent of cholic acid was included. Values are reported as the mean ± standard deviation for duplicate mea surements Substrate Detergent Specific activity (nmol/min/mg) DPPC DPPC+POPG DPPC TX-lOO TX -lOO TX-IOO+cholic acid 230±10 1070±30 510±50 46 L. J. Lefkowitz et al. increase in the hydrolysis of DPPC. To determine whether this activation might be due to surface charge, 5 mole percent of the anionic detergent cholic acid was included in a separate experiment.
4. Legend see p. 35 Human Group-V Phospholipase-A, Expression in Pichia pastoris 35 To simplify the purification of human group-V PLA2 from P. pa storis, we engineered an N-terminal His tag on the group- V gene; this tag could be removed by treatment with enterokinase. The removable His tag was constructed using a two-step PCR strategy, as shown in Fig. 3. In the first step of the PCR, the 5' portion of the gene was modified to include an enterokinase site using the 48-mer primer (CAC CAC CAC GAC GAT GAC GAT AAA GGC TTG CTG GAC CTA AAA TCA ATG) and a 3' primer that is complementary to a region of the pPIC9K vector located after the stop codon (CGA ATT AAT TCG CGG CCG CCC TAG GG).