BUSTER User Manual
Barnase Hard-NCS tutorial

Quick input guide: tutorial 3

BUSTER NCS-constrained refinement of a 25% incomplete structure

This tutorial has been run in the "Standard" user level. In this mode, default values are used for most of the input items. The ones that needed some extra attention are described below:

Input item	Notes
Partial Structure	The PDB file with the atoms whose xyzB's will be refined by BUSTER-TNT
Chemical composition	21x3=63 missing residues One additional oxygen atom for each of the 3x110 residues of the full protein (to mimic ordered water molecules) The missing atoms are given a slightly higher B value than the average B of the atoms in the partial model (<B>=14).
Missing atoms prior envelope	Uniform: this is the simplest choice, amounting to distributing the missing atoms anywhere but where the partial structure atoms are. If you wanted to improve on this, you could select: "From current phases": the current 2Fo-Fc map would be used to define the regions where the missing atoms are expected to be.
Solvent content	0.43%, as estimated from the nominal 3x110=330 residues in the a.u. of the P32 unit cell
Refinement: additional commands for the minimiser	RANGE B 1 300: no B factors will refine below 1; atoms whose B factor wants to refine above 300 will be rejected
TNT sequence file	Was prepared with the script $BDG_home/bin/buster/pirToSeq.pl starting from an ASCII 1-letter sequence file for the full protein
Weight on the x-rays residual	Set to 3.5 - a value low enough to keep a fairly tight stereochemistry.
TNT hard-NCS file	To constrain shifts for three chains to be the same, the NCS-operators are needed: CLUSTER PRO NCS A PROA B PROB C PROC TRANSFORMATION PROA SYSTEM TNT - MATRIX 1.00000 0.00000 0.00000 - 0.00000 1.00000 0.00000 - 0.00000 0.00000 1.00000 - SHIFT XYZ 0.0000 0.0000 0.0000 - SHIFT B 0.00 - SHIFT OCC 0.0000 TRANSFORMATION PROB SYSTEM TNT - MATRIX 0.94470 -0.31444 0.09308 - -0.31342 -0.94926 -0.02582 - 0.09648 -0.00478 -0.99532 - SHIFT XYZ 36.9543 62.2834 99.4107 - SHIFT B 0.02 - SHIFT OCC 0.0000 TRANSFORMATION PROC SYSTEM TNT - MATRIX 0.99173 -0.06937 0.10802 - -0.06771 -0.99753 -0.01892 - 0.10907 0.01145 -0.99397 - SHIFT XYZ 11.7003 30.7054 79.6683 - SHIFT B 0.21 - SHIFT OCC 0.0000 The NCS-operators were obtained by running TNT-ncs issuing the command GATHER.
Stereochemical restraints libraries	$BDG_tntdata/csdx_protgeo.dat: geometry of standard amino-acids $BDG_tntdata/bcorrel.dat: temperature factors correlation restraints for standard amino-acids $BDG_tntdata/contact.dat: basic non-bonded distances for most commonly occurring atom types