The conserved N-terminal heliz of acylpeptide hydrolase from archaeon Aeropyrum perniz K1 is important for its hyperthermophilic activity
The acylpeptide hydrolases from hyperthermophilic archaeon Aeropyrum pernix Kl has a short conserved N terminal helix in its family. The role of this N-terminal helix in the function of the hyperthermophilic enzyme, however, is unknown. Here, we investigated this question by protein engineering and biophysical methods. We found that a mutant (AN21) with the N-terminal helix deleted is no longer functional at the optimum temperature for WT enzyme (95℃), required for the survival of Aeropyrum pernix Kl. Instead, AN21 has the optimum activity at ～77℃, with higher activities than the WT enzyme below this temperature. AN21 is less stable than the WT enzyme and started unfolding at -77℃. indicating that the loss of the enzymatic activity of AN21 at higher temperature is due to its low thermodynamic stability. In addition, we found that the salt bridges formed between the N-terminal helix and the catalytic domain of the enzyme play only a minor role in stabilizing the enzyme, suggesting that hydrophobic interactions mainly contribute to the stabilization. Since the N-terminal helix is conserved in this family of enzymes, our results suggest that the N-terminal helix is likely to play an important role for stabilizing all other enzymes in this family.
Acylpeptide hydrolase Prolyl oligopeptidase family N-terminal truncation Protein stability Molecular dynamics simulation
Zuoming Zhang Baisong Zheng Yanping Wang Yiqian Chen Giuseppe Manco Yan Feng
Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, jitin Universi Key Laboratory for Molecular Enzymology and Engineering of the Ministry of Education, jitin Universi Institute of Protein Biochemistry, CNR, Via P.Castellino 111.80131 Napoli, Italy
国际会议
广州
英文
197-204
2008-11-01（万方平台首次上网日期，不代表论文的发表时间）