*Restraints* implies use of an energy function
without absolute fixing of the desired quantity; the refc
file can be used for restraining Cartesian coordinates of
selected atoms, or internal coordinate restraints can be
applied in Sander.

Here's an example of temperature control; note the "&rst iat=0, &end" that ends the section:

Run belly water at 300K to disorder wat216 periodicity then cool &cntrl irest=0, ntx=1, ibelly=1, imin=0, nrun=1, nstlim=2000, nsnb=25, tempi=10.0, temp0=300.0, ntt=1, npscal=1, ntc=2, ntf=2, cut=8.0, ntb=1, nmropt=1, iftres=0, &end # Warm up fast (actual temp will take a while) then after a while, cool down. # ATOM numbers need to be set for each system, e.g. 1st ion, last water atom. &wt type='TEMP0', istep1=1000, istep2=2000, value1=300.0, value2=10.0, iinc=5, &end &wt type='END', &end &rst iat=0, &end belly - waters, ions ATOM 531 9030 END END

*How to get cartesian and NMR restraints together?*

Put the nmr restraints into a DISANG file; they unfortunately cannot be in the mdin file itself.

Section Four of Sander input in the Amber 7 manual [Distance, angle,
and torsional restraints] describes how to restrain, not constrain,
dihedrals.
There is no way to keep torsion angles absolutely fixed
within AMBER. Incidentally, you can
easily create a *vs.* torsion
table using Interface.

Dave Case:

Yes. The variable IFVARI can be used to control individual restraint weights. See section 5.9 of the Amber 7 manual. For example, if you wanted to have a WC constraint on for the first 2500 steps, then ramp to zero during the next 2500 steps, you might do something like the following:

&rst iat=?,?, r1=???, r2=???, r3=???, r4=???, rk2=???, rk3=???, nstep1=0, nstep2=2500, &end &rst iat=?,?, nstep1=2501, nstep2=5000, ifvari=1, r1a=???, r2a=???, r3a=???, r4a=???, rk2a=0.0, rk3a=0.0, &endThe first &rst line sets up the constraint for the first 2500 steps, and it is constant (ifvari has the default value of zero.) The second &rst continues this constraint for another 2500 steps, but ramps the force constants rk2 and rk3 down to zero; as these get smaller, there essentially becomes no penalty for violating the constraint. After 5000 steps, neither constraint is active.

Refc cannot be used for internal (bond, angle or dihedral) constraints: it is for Cartesian restraints only. The format is the same as inpcrd and restrt.

R1, R2, R3, R4 define a flat-welled parabola which becomes linear beyond a specified distance. I.e.

\ / \ / \ / . . . . ._______. R1 R2 R3 R4 "\" = lower bound linear response region "/" = lower bound linear response region "." = parobola "_" = flat regionIf you have determined lower and upper bounds from an NMR experiment, those would typically correspond to R2 and R3. Note that the flat well means that any value R2 <= value <= R3 is equally acceptable. R1 and R4 define linear response regions. These are sometimes used so that restraints that severely violate the lower and upper bounds don't tear the structure up. A typical value of R1 is R2-2.0. And a typical value of R4 is R3+2.0 (angstroms).

Answer is in the manual, but somewhat hidden! See the discussion about the "IPNLTY" variable:

IPNLTY = 1 the program will minimize the sum of the absolute values of the errors; this is akin to minimizing the crystallographic R- factor (default). = 2 the program will optimize the sum of the squares of the errors. = 3 For NOESY intensities, the penalty will be of the form awt [Ic^(1/6)-Io^(1/6)]^2. Chemical shift penalties will be as for ipnlty=1.

Further discussion appears in section SIX, "Chemical Shift Restraints".

(5.11 page 119 in Amber7 manual)

Basically, the program will minimize the sum of the absolute values of the "errors", where in this case the "error" is the difference between the calculated and the observed shift. The SHRANG variable allows you to ignore errors less than some cutoff, and the WT variable allows you to weight some shifts more heavily than others in the sum.

Positional restraints work well enough to keep a molecule near its starting conformation, and they can force a molecule from one conformation to a second SIMILAR conformation. But they don't do a good job of forcing a molecule through substantial regions of conformational or distance space.

See src/nmr_aux/prepare_input/makeCHIR_RST.

This script will construct penalty functions to help keep all chiral carbons in the right places, even under high-T annealling.