Re: [AMBER] Help with Umbrella Sampling

From: bharat gupta <>
Date: Wed, 6 Jul 2016 14:31:57 +0900


I am studying a simultaneous proton transfer, bond breakage and nucleophilic
attack (by water molecule), using US approach for which I had already
performed 5ns QM/MM simulation.

All three reactions takes places in a single step (Inversion mechanism for
Glycoside hydrolase). Now, I am confused in defining the restraint
variables. Since I am new to US, it would be a great help if somebody can
guide me through this.

I have selected 4 Reaction Coordinates:

1. RC1: Proton transfer from Base residue to leaving group
     OE1-HE1 -> C----O4 (this glycosidic bond breaks and HE1 is transferred
to O4 )
    So, the reaction coordinate for this reaction is difference in distance
between OE1-HE1 and O4--HE1.

2. RC2: Glycosidic bond breakage:
    C-----O4 -> C O4. The reaction coordinate for this reaction is the
distance between C and O4

3. RC3: Nucleophilic attack by water:
     H(i)O(w)H(w) [this is nucleophilic water] ---- C (anomeric carbon of
the broken glycosidic bond). The reaction coordinate for this reaction is
the distance between C and O(w).

4. RC4: Proton transfer from water (H(i)) to Acid Residue
     H(i)O(w)H(w) -- OD1 (Acid residue). The reaction coordinate for this
step is difference in distance between O(w)-H(i) and OD1-H(i).

I also not able to understand how to make the restraint file for difference
in distances between two set of atoms, as in case of RC4 and RC1. I would
be helpful for me if somebody explains it too with an example.

For the RC2, I have made the following restraint file:
# distance restraint
 &rst iat=8122,8132 r1=0, r2=1.8, r3=1.8, r4=5, rstwt=1,-1, rk2 = 500.0,
rk3 = 500.0, /

I have increased the the value for r2 & r3 by 0.2 and upto 3.4. I am not
able to understand what should be the value for r1 and r4 ? Can anyone pls
comment on it and explain it briefly? The Simulation for the setting r2 &
r3 = 3.0 to 3.4 were unable to converge and failed with this error:

QMMM: Unable to achieve self consistency to the tolerances specified
QMMM: No convergence in SCF after 1000 steps.
QMMM: Job will continue with unconverged SCF. Warning energies
QMMM: and forces for this step will not be accurate.
QMMM: E = -0.2696E+07 DeltaE = 0.3931E+00 DeltaP = 0.3364E-01
QMMM: Smallest DeltaE = 0.3385E-06 DeltaP = 0.6093E-07 Step = 74

So, I tried to see the overlap for windows 1.8 to 2.8 and here is the

I am confused about choosing the number windows to sample for RC2. Should I
stop at r2 and r3 =2.8, as uptill here QM/MM simulation converges? Then in
that case I have only 6 windows only for RC2, is that sufficient enough to
calculate the free energy for bond breakge??

I checked at the structures from the window 3,4,5,6 and I found that the
bond is broken.

*Best Regards*
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Received on Tue Jul 05 2016 - 23:00:02 PDT
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