Tables for
Volume F
Crystallography of biological macromolecules
Edited by M. G. Rossmann and E. Arnold

International Tables for Crystallography (2006). Vol. F, ch. 25.2, p. 722   | 1 | 2 |

Section An example

V. S. Lamzin,n* A. Perrakiso and K. S. Wilsonp An example

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The structure of chitinase A from Serratia marcescens (Perrakis et al., 1994[link]) was initially solved by multiple isomorphous replacement with anomalous signal (MIRAS), with only a single derivative contributing to resolution higher than 5.0 Å. The MIRAS map (2.5 Å) was solvent-flattened. Model building was not straightforward and much time was spent in tracing the protein chain.

As an experiment, the solvent-flattened map was used to initiate building of free-atom models, using least-squares minimization against the native 2.3 Å data combined with ARP. This resulted in crystallographic R factors ranging between 20.1 and 22.4%. Each ARP model gave phases marginally worse than those available by solvent flattening alone, due to the limited resolution of the native data. However, the wARP averaging procedure resulted in a reduction of 11.2° in the weighted mean phase error. The map correlation coefficient between the final map and the wARP map was 81.2%, better by 12.8% compared with the solvent-flattened map.

The wARP model with the lowest R factor was used to initiate model building. In the initial tracing, 75 residues were identified, belonging to more than 20 different main-chain fragments. After autobuilding, ten cycles of restrained ARP were run according to the standard protocol. One REFMAC cycle of conjugate-gradient minimization was executed to optimize a maximum-likelihood residual and bulk solvent scaling. [\sigma_{A}]-weighted maps were calculated and ARP was used to update the model. All atoms (main-chain, side-chain and free atoms) were allowed to be removed and new atoms were added where appropriate. After ten iterations, a new building cycle was invoked. After every `big' cycle, a more complete model was obtained. This `big' cycle was iterated 20 times. Finally, 515 residues were traced in nine chains, all of which were docked unambiguously into the sequence. This is the lowest-resolution application to date. 2.3 Å was the real resolution limit of the data measured from these crystals; however, the high solvent content (61%) provided on average seven observations per atom and an almost complete trace was easily accomplished.


Perrakis, A., Tews, I., Dauter, Z., Oppenheim, A., Chet, I., Wilson, K. S. & Vorgias, C. E. (1994). Structure of a bacterial chitinase at 2.3 Å resolution. Structure, 2, 1169–1180.Google Scholar

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