Total direct chemical synthesis and biological activities of human group IIA secretory phospholipase A2.
Human group IIA secretory phospholipase A(2) (hGIIA sPLA(2)) is reported to be involved in inflammation, since its expression level is enhanced under various inflammatory conditions. In this work, we report the total chemical synthesis of this enzyme (124 amino acids) by solid-phase method. The identity of the protein, in denatured or folded (7 disulphide bonds) forms, was confirmed by electrospray MS. Synthetic sPLA(2) possesses the same circular dichroism spectrum, enzymic activity in hydrolysing different phospholipid substrates, and inhibitory effect in thrombin formation from prothrombinase complex as the recombinant sPLA(2). Furthermore, LY311727, a reported specific hGIIA sPLA(2) inhibitor, is able to inhibit the synthetic and the recombinant enzymes with the same efficiency. This study demonstrates that chemically continuous solid phase synthesis is an alternative and less time-consuming approach to producing small, structurally folded and fully active proteins of up to 124 amino acids, such as hGIIA sPLA(2). Moreover, this technique provides more flexibility in analogue synthesis to elucidate their physiological functions and pathological effects.
Dong
Chang-Zhi
CZ
Laboratoire de Pharmacochimie Moléculaire, Université Paris 7-Denis Diderot, Case 7066, 2, Place Jussieu, 75251 Paris Cedex 05, France.
Romieu
Anthony
A
Mounier
Carine M
CM
Heymans
Françoise
F
Roques
Bernard P
BP
Godfroid
Jean-Jacques
JJ
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D006801:Humans; D008969:Molecular Sequence Data; D010741:Phospholipases A / Q000138:chemical synthesis* / Q000378:metabolism / Q000494:pharmacology*; D054467:Phospholipases A2; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000138:chemical synthesis / Q000378:metabolism / Q000494:pharmacology; D021241:Spectrometry, Mass, Electrospray Ionization
D011994:Recombinant Proteins; D010741:Phospholipases A; D054467:Phospholipases A2