On Fri, Dec 05, 2025, cai.hanfeng--- via AMBER wrote:
>
>We are trying to prepare an rbLDH (rabbit lactate dehydrogenase) with
>ligands in 1 M trehalose (TRE), and have encountered an unexpected
>increase in box size after the parametrization step. This change alters
>the trehalose concentration from the intended 1 M to ~0.28 M. We would
>appreciate help understanding why the box expands and how to keep the
>concentration constant.
>
>Summary of our workflow:
>We first dissolved the complex [rbLDH + pyruvate (PYR) + NADH (NAI)]
>into TIP3P water, and parameterized the system in water. From the output
>topology and coordinate files, we extracted the box size (890,346 ų,
>truncated octahedron) and calculated the number of trehalose molecules
>that we need to add to achieve 1 M (need 537 TRE molecules). To prepare
>the system with trehalose, we first removed all water and ions from the
>original system, loaded the solute-only structure (rbLDH + PYR + NAI) into
>tleap, saved a PDB with a temporary box (90°), and then manually fixed
>CRYST1 to our original truncated octahedron. Then we used AddToBox tool
>to add 537 molecules of trehalose into the solute and kept the box size
>consistent. After that, we performed parameterization for the whole system
>(rbLDH + PYR + NAI + TRE), in which we also dissolved the system with TIP3P
>water. But after that we found the box was changed into a larger size
>(3.13922e+06 ų), and therefore the concentration of trehalose is no
>longer 1 M. We wonder why the box size will increase after the last step
>parameterization, or how to maintain the original box size, or how to keep
>the trehalose concentration constant after parameterization?
An increase of the volume size by a factor of three is indeed unusual. My
first guess is that you have way too much water in the final system, but you
should certainly visualize the starting and equilibrated structures to look
for possible voids.
How did you decide to add 53 waters via AddtoBox? And the solvateOct
command later on may set its own box: I'm not sure what what solvatOct with
a buffer distance of 0.0 will really do. If you want to add water to a
pre-built box with a given volume, you should probably use AddtoBox to do this.
You may have to determine the number of waters to add by trial and error, to
get a final system that stays close to the desired volume upon
equilibration.
...hope this helps....dac
_______________________________________________
AMBER mailing list
AMBER.ambermd.org
http://lists.ambermd.org/mailman/listinfo/amber
Received on Sat Dec 06 2025 - 10:30:05 PST
