%tleap

-I: Adding /usr/local/prog/amber16/dat/leap/prep to search path.

-I: Adding /usr/local/prog/amber16/dat/leap/lib to search path.
-I: Adding /usr/local/prog/amber16/dat/leap/parm to search path.
-I: Adding /usr/local/prog/amber16/dat/leap/cmd to search path.

Welcome to LEaP!
(no leaprc in search path)
>source leaprc.protein.ff14SB
en tleap

----- Source: /usr/local/prog/amber16/dat/leap/cmd/leaprc.protein.ff14SB

----- Source of /usr/local/prog/amber16/dat/leap/cmd/leaprc.protein.ff14SB done
Log file: ./leap.log
Loading parameters: /usr/local/prog/amber16/dat/leap/parm/parm10.dat
Reading title:
PARM99 + frcmod.ff99SB + frcmod.parmbsc0 + OL3 for RNA
Loading parameters: /usr/local/prog/amber16/dat/leap/parm/frcmod.ff14SB
Reading force field modification type file (frcmod)
Reading title:
ff14SB protein backbone and sidechain parameters
Loading library: /usr/local/prog/amber16/dat/leap/lib/amino12.lib
Loading library: /usr/local/prog/amber16/dat/leap/lib/aminoct12.lib
Loading library: /usr/local/prog/amber16/dat/leap/lib/aminont12.lib

laz = sequence {NGLN PHE GLY ALA GLU PRO VAL GLU GLY LYS PRO ILE CALA}
salvar la estructura en un fichero de libreria (lib) y como pdb
saveoff laz laz_lineal.lib
Creating laz_lineal.lib
Building topology.
Building atom parameters.

savepdb laz laz_lineal.pdb

Writing pdb file: laz_lineal.pdb
Converting N-terminal residue name to PDB format: NGLN -> GLN
Converting C-terminal residue name to PDB format: CALA -> ALA

Creación ficheros formato para uso en Amber

Hay que crear un fichero de parametros (.prmtop) y un fichero de coordenadas (.inpcrd)
 saveamberparm laz laz.top laz.crd

Checking Unit.
WARNING: The unperturbed charge of the unit: -0.999989 is not zero.

-- ignoring the warning.

Building topology.
Building atom parameters.
Building bond parameters.
Building angle parameters.
Building proper torsion parameters.
Building improper torsion parameters.
total 39 improper torsions applied
Building H-Bond parameters.
Incorporating Non-Bonded adjustments.
Not Marking per-residue atom chain types.
Marking per-residue atom chain types.
(Residues lacking connect0/connect1 -
these don't have chain types marked:

res total affected

CALA 1
NGLN 1
)
(no restraints)

Salimos de tleap
Quit

%ls –rtl
nos lista los ficheros que hemos generado
editamos pdb con vim o con joe
%vim laz_lineal.pdb

Pasamos el fichero al ordenador que usamos como terminal y lo abrimos usando pymol
Como hay problemas lo que vamos a realizar es una pequeña minimización de energía, para
relajar la molécula y que se separen las cadenas laterales.
Para ello creamos un fichero donde le vamos a decir que es lo que tiene que hacer:
mini_ini.in
Stage 1 - minimización del lazo
&cntrl
imin=1, maxcyc=1000, ncyc=500,
cut=16., rgbmax=16., igb=5, ntb=0,
ntpr=100, ntxo=1
/

imin Flag to run minimization.

= 0 (default) Run molecular dynamics without any minimization.
= 1 Perform an energy minimization

Energy minimization
maxcyc The maximum number of cycles of minimization. Default = 1.
ncyc If NTMIN is 1 then the method of minimization will be switched from steepest descent to conjugate
gradient after NCYC cycles. Default 10.

ntmin Flag for the method of minimization.

= 0 Full conjugate gradient minimization. The first 4 cycles are steepest descent at the start of
the run and after every nonbonded pairlist update.
= 1 For NCYC cycles the steepest descent method is used then conjugate gradient is switched on
(default).
= 2 Only the steepest descent method is used.
= 3 The XMIN method is used
= 4 The LMOD method is used

dx0 The initial step length. If the initial step length is too big then will give a huge energy; however
the minimizer is smart enough to adjust itself. Default 0.01.
drms The convergence criterion for the energy gradient: minimization will halt when the root-mean-
square of the Cartesian elements of the gradient is less than DRMS. Default 1.0E-4 kcal/mole-Å

igb Flag for using the generalized Born or Poisson-Boltzmann implicit solvent models. Default is 0.

cut This is used to specify the nonbonded cutoff, in Angstroms. For PME, the cutoff is used to limit
direct space sum, and 8.0 is usually a good value. When igb>0, the cutoff is used to truncate
nonbonded pairs (on an atom-by-atom basis); here a larger value than the default is generally
required. A separate parameter (RGBMAX) controls the maximum distance between atom pairs
that will be considered in carrying out the pairwise summation involved in calculating the
effective Born radii.
When igb > 0, the default is 9999.0 (effectively infinite)
When igb==0, the default is 8.0.

The Particle Mesh Ewald (PME) method is always "on", unless ntb = 0. PME is a fast implementation of
the Ewald summation method for calculating the full electrostatic energy of a unit cell.

ntb This variable controls whether or not periodic boundaries are imposed on the system during
the calculation of non-bonded interactions. Bonds spanning periodic boundaries are not yet
supported. There is no longer any need to set this variable, since it can be determined from igb
and ntp parameters. The “proper” default for ntb is chosen (ntb=0 when igb > 0, ntb=2 when
ntp > 0, and ntb=1 otherwise). This behavior can be overridden by supplying an explicit value,
although this is discouraged to prevent errors. The allowed values for NTB are
= 0 no periodicity is applied and PME is off (default when igb > 0)
= 1 constant volume (default when igb and ntp are both 0)
 = 2 constant pressure (default when ntp > 0)
Constant pressure is not used in minimization (IMIN=1)

ntpr Every ntpr steps, energy information will be printed in human-readable form to files "mdout"
and "mdinfo". "mdinfo" is closed and reopened each time, so it always contains the most
recent energy and temperature. Default 50.

ntxo Format of the final coordinates, velocities, and box size (if constant volume or pressure run)
written to file "restrt".
= 1 Formatted (ASCII)
= 2 (default) NetCDF file (recommended, unless you have a workflow that requires the formatted
form.)
Una vez creado el fichero introducimos la orden de ejecución de la simulación:

%sander -O -i mini_ini.in -o mini_ini.out -p laz.top -c laz.crd -r mini_ini.rst

SANDER
sander [-help] [-O] [-A] -i mdin -o mdout -p prmtop -c inpcrd -r restrt
-ref refc -mtmd mtmd -x mdcrd -y inptraj -v mdvel -frc mdfrc -e mden
-inf mdinfo -radii radii -cpin cpin -cpout cpout -cprestrt cprestrt
-evbin evbin -suffix suffix
-O Overwrite output files if they exist.
-A Append output files if they exist (used mainly for replica exchange)

Las opciones son resumidamente:

mdin input control data for the min/md run

mdout output user readable state info and diagnostics -o stdout will send output to stdout (to
the terminal) instead of to a file.
mdinfo output latest mdout-format energy info
prmtop input molecular topology, force field, periodic box type, atom and residue names
inpcrd input initial coordinates and (optionally) velocities and periodic box size
refc input (optional) reference coords for position restraints; also used for targeted MD
mtmd input (optional) containing list of files and parameters for targeted MD to multiple
targets
mdcrd output coordinate sets saved over trajectory
inptraj input coordinate sets in trajectory format, when imin=5
mdvel output velocity sets saved over trajectory
mdfrc output force sets saved over trajectory
mden output extensive energy data over trajectory (not synchronized with mdcrd or mdvel)
restrt output final coordinates, velocity, and box dimensions if any - for restarting run
inpdip I nput polarizable dipole file, when indmeth=3
rstdip output polarizable dipole file, when indmeth=3
cpin input protonation state definitions
cprestrt protonation state definitions, final protonation states for restart (same format as cpin)
cpout output protonation state data saved over trajectory
evbin input input for EVB potentials
suffix output this string will be added to all unspecified output files that are printed (for
multisander runs, it will append this suffix to all output files)
NSTEP ENERGY RMS GMAX NAME NUMBER
1000 -3.5759E+02 4.8786E-01 2.6707E+00 OXT 133

BOND = 7.0915 ANGLE = 27.1176 DIHED = 118.1621

VDWAALS = -25.7290 EEL = -942.9471 EGB = -340.0875
1-4 VDW = 30.4734 1-4 EEL = 768.3252 RESTRAINT = 0.0000
Extraemos la estructura minimizada en format pdb para poder manipularla y verla en pymol

%ambpdb -p laz.top –c mini_ini.rst > mini_ini.pdb

Ejercicio 2
De la estructura obtenida en pdb, la cargamos en pymol, le añadimos hidrógenos y la salvamos,
tendremos en este momento 190 átomos.
Este segundo ejercicio consiste en usar métodos semiempíricos para la optimización de la geometría.
Para ello vamos a utilizar el programa GAUSSIAN 09, utilizando como método semiempírico
PM3

Lo primero que hay que realizar es una conversión del fichero pdb a formato xyz, utilizamos el programa
newzmat

http://www.gaussian.com/g_tech/g_ur/u_newzmat.htm

newzmat –ipdb mini_ini.pdb –oxyz min3.xyz

>carga multiplicidad [0,1]? -1,0

editamos el fichero generado y le añadimos las líneas de comando del gaussian

vim min3.xyz

nohup g09 <<END >lazmin.log

#PM3 Opt

-1 1
N 3.281 1.608 0.333
C 3.976 2.919 0.245
C 5.528 2.787 0.303
O 6.035 1.805 0.855
C 3.443 3.834 1.367
C 3.725 5.333 1.178
C 3.228 6.193 2.345

N 4.108 6.937 2.997

O 2.046 6.225 2.681
H 3.711 3.379 -0.710
H 3.784 7.512 3.761
H 5.084 6.947 2.731
H 3.851 0.910 0.807
….

La geometría obtenida la podemos observar con el molden.exe.

Ejercicio 2-bis

Efecto del cut en los cálculos de MD

Vamos a generar una molécula de dsDNA poly-A
Utilizamos el generador nab de amber

Creamos un fichero con vim:

vim dsdna.nab

molecule m;

m = fd_helix( "abdna", "aaaaaaaaaa", "dna" );

putpdb( "dsdna.pdb", m, "-wwpdb");

ejecutar nab

nab dsdna.nab

./a.out

Cargamos en tleap la estructura generada en pdb

>tleap

>source leaprc.DNA.OL15

>source leaprc.water.tip3p

dna=loadpdb “dsdna.pdb”

Creamos los ficheros de coordenadas y parámetros

saveamberparm dna dsdna.top dsdna.crd

Realizamos una minimización y dinámica molecular en vacio

Minimización

Generamos un fichero min.in

Vim mindna.in

polyA-polyT 10-mer: minimización inicial prior to MD

&cntrl
imin = 1,
maxcyc = 500,
ncyc = 250,
ntb = 0, ntx0 =1,
igb = 0,
 cut = 12
/

sander –O –i mind.in –o dsdna_min.out –c dsdna.crd –p dsdna.top –r dsdna_min.rst

Ahora vamos a lanzar la simulación de MD, en dos situaciones distintas, una es con un cut=12A y la otra
con cut=999

Sólo vamos a analizar los primeros 100ps por tanto la simulación se ceñirá a estos.

Generamos los ficheros de coordenadas y topología a partir de los minimizados

Generamos el fichero input

Vim dsdna_12Acut.in

10-mer DNA MD en-vacio, 12 A cut off

&cntrl
imin = 0, ntb = 0,
igb = 0, ntpr = 100, ntxo = 1, ioutfm=0, ntwx = 100,
ntt = 3, gamma_ln = 1.0,
tempi = 300.0, temp0 = 300.0
nstlim = 100000, dt = 0.001,
cut = 12.0
/

Esta simulación es a 300ºK para mantener la temperatura se activa la variable ntt y se le dá el valor 3,
con esta asignación el método que se utiliza para mantener la temperatura en equilibrio es el método
dinámico de langevin con una frecuencia de colisión dada por la variable gamma_ln
Ntb e igb son igual a cero.
Nstlim=100000, se van a realizar 100000 pasos de simulación con un tiempo para cada paso de 1fs
dt=0.001 lo que supone 100ps (100000x1fs)

Igualmente generamos el fichero input para cuando no hay cut

Vim dsdna_nocut.in

10-mer DNA MD en-vacio, 12 A cut off

&cntrl
imin = 0, ntb = 0,
igb = 0, ntpr = 100, ntxo =1, ioutfm=0, ntwx = 100,
ntt = 3, gamma_ln = 1.0,
tempi = 300.0, temp0 = 300.0
nstlim = 100000, dt = 0.001,
cut = 999
/

Ejecutamos las simulaciones

Sander –O –i dsdna_12Acut.in –o dsdna_12Ac.out –c dsdna_min.rst –p dsdna.top –r dsdna_12Ac.rst –x

dsdna_12Ac.mdcrd

Sander –O –i dsdna_nocut.in –o dsdna_nc.out –c dsdna_min.rst –p dsdna.top –r dsdna_nc.rst –x

dsdna_nc.mdcrd
Ejercicio 3
La siguiente fase es el calentamiento del sistema. Realizaríamos en una aproximación de
dinámica molecular (MD) sobre el Sistema. Lo primero que vamos a hacer es calentar
el sistema lentamente pasando desde 0k a 300k (temperatura ambiente). Este
calentamiento lo llevaremos a cabo en 7 fases de 50 ps cada una, en cada fase se
aumentará la temperatura 50k. Por qué hacerlo así, es para que se reduzcan los
cambios en el Sistema debido al calentamiento brusco, permitiendo que se estabilice
en cada una de las fases.

Usando la geometría optimizada por el gaussian vamos a calentar el sistema de 0ºK a 300ºK.
Para cargar la geometría obtenida en gaussian usamos el programa Antechamber
Antechamber –L nos da una ayuda sobre los tipos de ficheros que maneja
Antechamber –help nos da una ayuda
Una vez que generamos el fichero en un formato mol2, ya que es el formato que se necesita para el
siguiente programa parmchk2, que nos genera un fichero en formato frcmod
De parámetros que incluye la modificación del campo de fuerza necesaria para leap.

calor1.in
Fase 1 Calentamiento de laz2 0 to 50K
&cntrl
imin=0, irest=0, ntx=1,
nstlim=10000, dt=0.0005,
ntc=2, ntf=2,
ntt=1, tautp=1.0,
tempi=0.0, temp0=50.0,
ntpr=50, ntwx=50, ntxo=1, ioutfm=0,
ntb=0, igb=5,
cut=16.,rgbmax=16.
/

irest Flag to restart a simulation.

= 0 (default) Do not restart the simulation; instead, run as a new simulation. Velocities in the
input coordinate file, if any, will be ignored, and the time step count will be set to 0

= 1 Restart the simulation, reading coordinates and velocities from a previously saved restart
file. The velocity information is necessary when restarting, so ntx (see above) must be 4 or
higher if irest= 1

nstlim Number of MD-steps to be performed. Default 1.

dt The time step (psec). Recommended MAXIMUM is .002 if SHAKE is used, or .001 if it isn’t. Note
that for temperatures above 300K, the step size should be reduced since greater temperatures mean
increased velocities and longer distance traveled between each force evaluation, which can lead to
anomalously high energies and system blowup. Default 0.001.

ntt Switch for temperature scaling. Note that setting ntt=0 corresponds to the microcanonical (NVE) ensemble
(which should approach the canonical one for large numbers of degrees of freedom). Some aspects of the "weak-
coupling ensemble" (ntt=1) have been examined, and roughly interpolate between the microcanonical and
canonical ensembles.[294, 295] The ntt=2 and 3 options correspond to the canonical (constant T) ensemble.
= 0 Constant total energy classical dynamics (assuming that ntb<2, as should probably always be
the case when ntt=0).
 = 1 Constant temperature, using the weak-coupling algorithm.[296] A single scaling factor is
used for all atoms. Note that this algorithm just ensures that the total kinetic energy is
appropriate for the desired temperature; it does nothing to ensure that the temperature is even
over all parts of the molecule. Atomic collisions will tend to ensure an even temperature
distribution, but this is not guaranteed, and there are many subtle problems that can arise with
weak temperature coupling.[297] Using ntt=1 is especially dangerous for generalized Born
simulations, where there are no collisions with solvent to aid in thermalization.) Other
temperature coupling options (especially ntt=3) should be used instead.
= 2 Andersen temperature coupling scheme,[298] in which imaginary "collisions" randomize the
velocities to a distribution corresponding to temp0 every vrand steps. Note that in between
these "massive collisions", the dynamics is Newtonian. Hence, time correlation functions (etc.)
can be computed in these sections, and the results averaged over an initial canonical
distribution. Note also that too high a collision rate (too small a value of vrand) will slow down
the speed at which the molecules explore configuration space, whereas too low a rate means
that the canonical distribution of energies will be sampled slowly. A discussion of this rate is
given by Andersen.[299]
= 3 Use Langevin dynamics with the collision frequency g given by gamma_ln, discussed below.
Note that when g has its default value of zero, this is the same as setting ntt = 0. Since Langevin
simulations are highly susceptible to "synchronization" artifacts,[300, 301] you should explicitly
set the ig variable (described below) to a different value at each restart of a given simulation.
ntxo Format of the final coordinates, velocities, and box size (if constant volume or pressure run) written
to file "restrt".
= 1 Formatted (ASCII)
= 2 (default) NetCDF file (recommended, unless you have a workflow that requires the
formatted form.)

ntpr Every ntpr steps, energy information will be printed in human-readable form to files "mdout" and
"mdinfo". "mdinfo" is closed and reopened each time, so it always contains the most recent energy and
temperature. Default 50.

ntwx Every ntwx steps, the coordinates will be written to the mdcrd file. If ntwx = 0, no coordinate
trajectory file will be written. Default = 0.

ioutfm The format of coordinate and velocity trajectory files (mdcrd, mdvel and inptraj). As of
Amber 9, the binary format used in previous versions is no longer supported; binary
output is now in NetCDF trajectory format. Binary trajectory files have many
advantages: they are smaller, higher precision, much faster to read and write, and able
to accept a wider range of coordinate (or velocity) values than formatted trajectory files.
= 0 Formatted ASCII trajectory
= 1 (default) Binary NetCDF trajectory

temp0 Reference temperature at which the system is to be kept, if ntt > 0. Note that for temperatures
above 300K, the step size should be reduced since increased distance traveled between evaluations can
lead to SHAKE and other problems. Default 300.

tautp Time constant, in ps, for heat bath coupling for the system, if ntt = 1. Default is 1.0. Generally,
values for TAUTP should be in the range of 0.5-5.0 ps, with a smaller value providing tighter coupling to
the heat bath and, thus, faster heating and a less natural trajectory. Smaller values of TAUTP result in
smaller fluctuations in kinetic energy, but larger fluctuations in the total energy. Values much larger
than the length of the simulation result in a return to constant energy conditions.

ntc Flag for SHAKE to perform bond length constraints.[306] (See also NTF in the Potential function
section. In particular, typically NTF = NTC.) The SHAKE option should be used for most MD calculations.
The size of the MD timestep is determined by the fastest motions in the system. SHAKE
removes the bond stretching freedom, which is the fastest motion, and consequently allows a larger
timestep to be used. For water models, a special "three-point" algorithm is used.[307] Consequently, to
employ TIP3P set NTF = NTC = 2.
Since SHAKE is an algorithm based on dynamics, the minimizer is not aware of what SHAKE is
doing; for this reason, minimizations generally should be carried out without SHAKE. One exception is
short minimizations whose purpose is to remove bad contacts before dynamics can begin.
For parallel versions of sander only intramolecular atoms can be constrained. Thus, such atoms must be
in the same chain of the originating PDB file.
= 1 SHAKE is not performed (default)
= 2 bonds involving hydrogen are constrained
= 3 all bonds are constrained (not available for parallel or qmmm runs in sander)

calor2.in
Stage 1 Calentamiento de laz2 50K to 100K
&cntrl
imin=0, irest=1, ntx=5,
nstlim=10000, dt=0.0005,
ntc=2, ntf=2,
ntt=1, tautp=1.0,
tempi=50.0, temp0=100.0,
ntpr=50, ntwx=50, ntxo=1, ioutfm=0,
ntb=0, igb=5,
cut=16., rgbmax=16.
/

……

Una vez escritos los siete ficheros para el calentamiento, preparamos un fichero para lanzarlo

Calentar

$AMBERHOME/bin/sander -O -i calor1.in -p laz2.prmtop -c

mini_ini.rst -r calor1.rst -o calor1.out -x calor1.mdcrd
echo "Alcanzados 50K"
gzip -9 calor1.mdcrd
$AMBERHOME/bin/sander -O -i calor2.in -p laz2.prmtop -c calor1.rst
-r calor2.rst -o calor2.out -x calor2.mdcrd
echo "Alcanzados 100K"
gzip -9 calor2.mdcrd
$AMBERHOME/bin/sander -O -i calor3.in -p laz2.prmtop -c calor2.rst
-r calor3.rst -o calor3.out -x calor3.mdcrd
echo "Alcanzados 150K"
gzip -9 calor3.mdcrd
$AMBERHOME/bin/sander -O -i calor4.in -p laz2.prmtop -c calor3.rst
-r calor4.rst -o calor4.out -x calor4.mdcrd
echo "Alcanzados 200K"
gzip -9 calor4.mdcrd
$AMBERHOME/bin/sander -O -i calor5.in -p laz2.prmtop -c calor4.rst
-r calor5.rst -o calor5.out -x calor5.mdcrd
echo "Alcanzados 250K"
gzip -9 calor5.mdcrd
$AMBERHOME/bin/sander -O -i calor6.in -p laz2.prmtop -c calor5.rst
-r calor6.rst -o calor6.out -x calor6.mdcrd
gzip -9 calor6.mdcrd
echo "Alcanzados 300K"

lo salvamos y lo hacemos ejecutable

Para observar la dinámica molecular, los ficheros *.mdcrd utilizamos el VMD

Ahora extraemos de los fichero .out las energías y las representamos.
Un ejemplo de comando para la extracción de EKtot es:

grep “EKtot” calor*.out

genera en pantalla la siguiente salida

…..
calor4.out: Etot = -238.6200 EKtot = 69.3072 EPtot = -307.9273
…….
calor6.out: Etot = -163.8971 EKtot = 112.2405 EPtot = -276.1375
…..
Con el commando

grep “EKtot” calor*.out | awk '{print $5,$6,$7}' > ektot.txt

obtendriamos en un fichero

……
EKtot = 115.9015
EKtot = 103.4956
EKtot = 104.0817
EKtot = 113.9150
EKtot = 103.3006
EKtot = 99.6208
……
Ejercicio 4
Le añadimos al Sistema agua

Lo vamos a realizar sobre la molécula sin plegar y luego la optimizada con pm3

Si quisiéramos utilizar cualquier otra estructura para este ejercicio lo que haríamos es pasar el fichero a
pdb con el comando

%ambpdb -p laz.top –c mini_ini.rst > mini_ini.pdb

y cargarla en tleap para introducirle el agua

%tleap
>source leaprc.protein.ff14SB
>source leaprc.water.tip3p

solvateBox solute solvent distance [ closeness ]

solvateOct solute solvent distance [ closeness ]

The solvateBox command creates a periodic solvent rectangular box around the solute UNIT.
The shape for solvateOct is a truncated octahedron. The solute UNIT is modified by the addition
of solvent RESIDUEs, such that the closest distance between any atom of the solute and the
edge of the periodic box is given by the distance parameter. The solvent box will be repeated
in all three spatial directions.
The optional closeness parameter can be used to control how close, in angstroms, solvent
ATOMs can come to solute ATOMs. The default value of the closeness argument is 1.0.
Smaller values allow solvent ATOMs to come closer to solute ATOMs. The criterion for
rejection of overlapping solvent RESIDUEs is if the distance between any solvent ATOM to
the closest solute ATOM is less than the sum of the ATOMs VANDERWAAL’s distances
multiplied by the closeness argument.
> mol = loadpdb my.pdb
> solvateOct mol TIP3PBOX 12.0 0.75

solvateCap solute solvent position radius [ closeness ]

The solvateCap command creates a solvent cap around the solute UNIT. The solute UNIT is
modified by the addition of solvent RESIDUEs. The solvent box will be repeated in all three
spatial directions to create a large solvent sphere with a radius of radius angstroms.
The position argument defines where the center of the solvent cap is to be placed. If position
is a RESIDUE, ATOM, or a LIST of UNITs, RESIDUEs, or ATOMs, then the geometric center
of the ATOMs within the object will be used as the center of the solvent cap sphere. If position
is a LIST containing three NUMBERS, then the position argument will be treated as a vector
that defines the position of the solvent cap sphere center.
The optional closeness parameter can be used to control how close, in angstroms, solvent
ATOMs can come to solute ATOMs. The default value of the closeness argument is 1.0. Smaller
values allow solvent ATOMs to come closer to solute ATOMs. The criterion for rejection of
63
3 LEaP
overlapping solvent RESIDUEs is if the distance between any solvent ATOM to the closest
solute ATOM is less than the sum of the ATOMs VANDERWAAL’s distances multiplied by the
closeness argument.
This command modifies the solute UNIT in several ways. First, the UNIT is modified by the
addition of solvent RESIDUEs copied from the solvent UNIT. Secondly, the cap parameter of
the UNIT solute is modified to reflect the fact that a solvent cap has been created around the
solute.
> mol = loadpdb my.pdb
> solvateCap mol WATBOX216 mol.2.CA 12.0 0.75

solvateShell solute solvent thickness [ closeness ]

The solvateShell command adds a solvent shell to the solute UNIT. The resulting
solute/solvent UNIT will be irregular in shape since it will reflect the contours of the solute.
The solute UNIT is modified by the addition of solvent RESIDUEs. The solvent box will be
repeated in three directions to create a large solvent box that can contain the entire solute and a
shell thickness angstroms thick. The solvent RESIDUEs are then added to the solute UNIT if
they lie within the shell defined by thickness and do not overlap with the solute ATOMs. The
optional closeness parameter can be used to control how close solvent ATOMs can come to
solute ATOMs. The default value of the closeness argument is 1.0. Please see the solvateBox
command for more details on the closeness parameter.
> mol = loadpdb my.pdb
> solvateShell mol WATBOX216 12.0 0.8

>laz=loadpdb "lazop3_1.pdb"
Loading PDB file: ./lazop3_1.pdb
total atoms in file: 133

> solvateOct laz TIP3PBOX 10 0.75

Scaling up box by a factor of 1.268708 to meet diagonal cut criterion
Solute vdw bounding box: 30.024 20.438 14.900
Total bounding box for atom centers: 55.399 55.399 55.399
(box expansion for 'iso' is 66.3%)
Solvent unit box: 18.774 18.774 18.774
Volume: 89696.658 A^3 (oct)
Total mass 47246.280 amu, Density 0.875 g/cc
Added 2548 residues.

> saveamberparm laz lazop3_w.top lazop3_w.crd

Checking Unit.
WARNING: The unperturbed charge of the unit: -0.999989 is not zero.

-- ignoring the warning.

Building topology.
Building atom parameters.
Building bond parameters.
Building angle parameters.
Building proper torsion parameters.
Building improper torsion parameters.
total 39 improper torsions applied
Building H-Bond parameters.
Incorporating Non-Bonded adjustments.
Not Marking per-residue atom chain types.
Marking per-residue atom chain types.
(Residues lacking connect0/connect1 -
these don't have chain types marked:

res total affected

CALA 1
NGLN 1
WAT 2548
)
(no restraints)

Lo pasamos a pdb para verlo

%ambpdb -p lazop3_w.top –c lazop3_w.crd > lazop3_w.pdb

Calentamos nuevamente pero con el agua.

Calentar_agua

$AMBERHOME/bin/sander -O -i calor1.in -p lazop3_w.prmtop -c

lazop3_w.pdb -r calor1w.rst -o calor1w.out -x calor1w.mdcrd
echo "Alcanzados 50K con agua"
$AMBERHOME/bin/sander -O -i calor2.in -p laz2.prmtop -c calor1.rst
-r calor2.rst -o calor2.out -x calor2.mdcrd
echo "Alcanzados 100K"
gzip -9 calor2.mdcrd
$AMBERHOME/bin/sander -O -i calor3.in -p laz2.prmtop -c calor2.rst
-r calor3.rst -o calor3.out -x calor3.mdcrd
echo "Alcanzados 150K"
gzip -9 calor3.mdcrd
$AMBERHOME/bin/sander -O -i calor4.in -p laz2.prmtop -c calor3.rst
-r calor4.rst -o calor4.out -x calor4.mdcrd
echo "Alcanzados 200K"
gzip -9 calor4.mdcrd
$AMBERHOME/bin/sander -O -i calor5.in -p laz2.prmtop -c calor4.rst
-r calor5.rst -o calor5.out -x calor5.mdcrd
echo "Alcanzados 250K"
gzip -9 calor5.mdcrd
$AMBERHOME/bin/sander -O -i calor6.in -p laz2.prmtop -c calor5.rst
-r calor6.rst -o calor6.out -x calor6.mdcrd
gzip -9 calor6.mdcrd
echo "Alcanzados 300K"

y comparamos

para extraer del fichero los datos podemos utilizar la siguiente orden:

grep "EKtot" calor?.out | awk '{print $7}'>ektot.txt

grep "EKtot" calor*w.out | awk '{print $7}' > ektotw.txt

Ejercicio 5 Docking

Se van a utilizar los programas Autodock vina, MGLTools y amber para realizar el
docking entre seroalbumina y palmitato.

Entramos en la base de datos de estructura pdb, buscamos la estructura

correspondiente a seroalbumina (1ao6.pdb), la descargamos en nuestro ordenador,
comprobamos que no tengan algunos de los aminoácidos conformaciones distintas, si
las tuvieran nos quedaríamos con la conformación A.
En amber vamos a construir la estructura del palmitato.
%tleap

>source leaprc.lipid17
>li=PA
>savepdb li palmitoil.pdb
>quit
Una vez fuera del tleap, le añadimos los hidrógenos al palmitoil.pdb
Usamos el comando reduce de amber
%reduce palmitoil.pdb >palmitoil_h.pdb

Entramos en el programa autodock de MGTools

En el menú File > Read Molecule

Edit > Hidrogens > add
All Hydrogens
noBondOrder
yes
En el menu Grid > Grid Box
Establece una caja, que podemos desplazar a la zona de interacción que nos interesa
o podemos hacerla crecer hasta incluir toda la proteína si nos interesa.
En nuestro caso vamos a situarla en las coordenadas del centro
X=25
Y=8.39
Z=23.3
Con un tamaño de puntos en la dimensión
X=110
Y=126
Z=104
Y un espaciado en angstrom de 0.792

En el menú Grid >Macromolecule >choose

Nos dirá las correcciones que realiza y abrirá un menú para guardar los cambios
y la molécula con formato .pdbqt
Lo salvamos como albu_h1.pdbqt
En el menú Ligand > input
Palmitoil_h.pdb
Add gasteiger charges…..
Menú Ligand > output > Save as PDBQT
Palmitoil_h1.pdbqt

Salimos del adt

En el servidor generamos un fichero de configuración que vamos a usar para lanzar el

autodock vina

% vim conf1
receptor= albu_h1.pdbqt
ligand= palmitoil_h1.pdbqt

center_x= 25
center_y= 8.4
center_z= 23.3

size_x= nºde puntos x 0.79 Å

size_y= 99.54
size_z= 85.32

exhaustiveness= 8

salir

% nohup vina --config conf1 -- out albu_pal.pdbqt -- log albu_pal.log &

#################################################################
# If you used AutoDock Vina in your work, please cite: #
# #
# O. Trott, A. J. Olson, #
# AutoDock Vina: improving the speed and accuracy of docking #
# with a new scoring function, efficient optimization and #
# multithreading, Journal of Computational Chemistry 31 (2010) #
# 455-461 #
# #
# DOI 10.1002/jcc.21334 #
# #
# Please see http://vina.scripps.edu for more information. #
#################################################################

WARNING: The search space volume > 27000 Angstrom^3 (See FAQ)
Detected 2 CPUs
Reading input ... done.
Setting up the scoring function ... done.
Analyzing the binding site ... done.
Using random seed: 1380633772
Performing search ... done.
Posteriormente analizamos los resultados con la aplicación autodock de mgltools