Questions and problems?

Constraints and Restraints

Nomenclature: the terms `constraints' and `restraints' are often used interchangeably. Constraints implies absolutely fixed values - the only ways to achieve these generally within AMBER is to use the `belly' option (Cartesian coordinates) or SHAKE (bond lengths). [Gibbs also allows bond, angle and dihedral constraints - see INTR in the Gibbs manual or read the Interface/AMBER manual.]

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.

How to get belly and NMR restraints together

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,

# 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

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.

... how to constrain a dihedral

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 energy vs. torsion table using Interface.

Is it possible to operate two independent sets of distance angle restraints, allowing both to be initially ramped in and then after 20-25ps the [e.g.] Watson/Crick distance and angle restraints to be slowly removed leaving the NMR derived restraints in place ??

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,   &end
The 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.
How can we make the file 'refc' for running with internal constraints?

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.

Please explain distance restraints in sander.

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 region

If 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).
What is the form of the potential used for chemical shift restraints?

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

                 = 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

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.

How about using positional restraints to force conformational changes?

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.

Is there a way Amber/Sander rtMD simulation can be restrained so that D amino acids are forbidden even though I have bad NMR restraints?

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.