title: Tryptophan 140 is important, but serine 141 Is essential for the formation of the integrated conformation of staphylococcal nuclease.
abstract: A series of N-terminal fragments of staphylococcal nuclease with different chain lengths has been taken as an in vitro nascent peptide folding model. Previous studies have shown that nascent peptide folding of the nuclease may begin early in the synthetic process with the content of ordered secondary structure increases with increasing peptide chain length, and that conformational adjustments are observed at certain stages during nascent peptide folding. Here, we focus attention on the conformational changes in the later stage of nascent peptide folding of the nuclease when the N-terminal fragment elongates nearly to the C-terminus of the nuclease in order to determine the role of the C-terminal region of the nuclease in the formation of the integrated conformation of the nuclease. We compared the conformational features of SNase R and its larger N-terminal fragments SNR135, SNR139, SNR140, and SNR141 using circular dichroism spectra, ANS-binding fluorescence and intrinsic fluorescence spectra. The results show that Trp140 is important for the enrichment of ordered secondary structure and for producing a greater ability to fold into a native-like conformation, but Ser141 is essential for the formation of the integrated conformation of the nuclease with a tightly packed tertiary structure. Note that the addition of only one residue to the C-terminus of elongating peptide chain can cause a dramatic change in conformation. The data also show the occurrence of continuous adjustments in conformation during peptide elongation, even after a rigid tertiary structure has formed, suggesting that the last eight residues (residues 142-149), which are disordered at the C-terminus of the nuclease, also possess a structural role, forming the native tertiary structure to provide a framework for the active site, even though they are remote from the active site in both sequence and spatial structure.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Academia Sinica, Beijing 100101, People's Republic of China.
authors.forename: J
authors.initials: J
authors.lastname: Yin
authors.forename: G
authors.initials: G
authors.lastname: Jing
mesh_terms: D000817:Anilino Naphthalenesulfonates / Q000737:chemistry; D002942:Circular Dichroism; D005456:Fluorescent Dyes / Q000737:chemistry; D019791:Guanidine / Q000737:chemistry; D008836:Micrococcal Nuclease / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012694:Serine; D013050:Spectrometry, Fluorescence; D014364:Tryptophan
chemical_list: D000817:Anilino Naphthalenesulfonates; D005456:Fluorescent Dyes; D010446:Peptide Fragments; D011994:Recombinant Proteins; D012694:Serine; C027132:1-anilino-8-naphthalenesulfonate; D014364:Tryptophan; C104574:staphylococcal nuclease R; D008836:Micrococcal Nuclease; D019791:Guanidine