It is worth mentioning that sPBP6, which is the next nearest homo

It is worth mentioning that sPBP6, which is the next nearest homolog of DacD, is inactive on pentapeptide substrate (Chowdhury et al., 2010). The crystal structures of sPBP5 and sPBP6 (Nicholas et al., 2003; Chen et al., 2009) show a similar secondary structure with no gross architectural differences. In the absence of crystal structure, CD spectral analysis would be of utmost importance to elucidate the biophysical characteristics of sDacD. It was evident from the CD spectra that purified protein was in a native conformation with characteristics of the molecular

spectra http://www.selleckchem.com/products/XL184.html of alpha and beta structures, indicating the protein was active and stable at room temperature. Unlike sPBP5 and sPBP6 (Chowdhury et al., 2010), more beta-sheets were detected in sDacD (Table 3, Supporting Information, Fig. S1). The occurrence of a larger amount of β-sheet structure in sDacD may cause some structural alteration, which might exert different biological activity than PBP5.

Because DacD shared a high level of aa identity with PBP5, homology modelling (or comparative protein structure modelling) could be applied to generate the three-dimensional conformation of sDacD. For model building, the program modeller 9v1 was used with the pdb coordinate, 3BEC chain A (crystal structure of E. coli PBP5 in complex with a peptide-mimetic cephalosporin; Sauvage et al., 2008) as template. The secondary structure prediction by Silmitasertib in vivo predict protein and psipred suggested that sDacD was a αβ protein with a larger amount of β-sheet structure (Table 3 and Fig. S2), which was consistent with the results obtained from CD spectroscopic analyses. The model of lowest energy value had 94.9% residues in the most favoured region in the Ramachandran plot and 98.35% residues had an average Adenosine triphosphate 3D-1D score above 0.2, as obtained through verify3d profile (Fig. 2), which affirms a well derived model. The model has been

deposited to the PMDB server (ID PM0076504). Like sPBP5, the sDacD model is composed of two Domains placed perpendicular to each other. Domain II is β-sheet-rich, whereas Domain I is composed of both α-helices and β-sheets (Fig. 2a). There is a relative increase in beta-sheet in Domain I of sDacD as compared with sPBP5. Comparison of the calculated secondary structure of the sDacD model generated by stride with that of sPBP5 indicates that residues Gln 38-Arg 39 and His158-Ser159-Ser160 of sDacD create a beta-sheet structure, whereas the respective positions of sPBP5 create coils and turns. Moreover, the Glu 230 and Met 233 of sPBP5 Domain I form turns, whereas the corresponding residues (Gln 229 and Arg 232, respectively) in the sDacD model adopt a beta-conformation. Therefore, both similarities and the differences exist when we take a closer view at the active-site of sDacD and sPBP5.

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