The first thing you want to do when debugging hydrogens in refmac5 is to
make sure it outputs whatever it was doing:
MAKE HOUT Y
This is not the default. Normally, refmac will take a PDB file without
any hydrogens in it, quietly add them before doing refinement, and then
delete them before outputting the final coordinates. Why so secretive?
Probably because someone complained about having to look at all that
fuzzy hydrogen nonsense in the graphics. In fact, I know some
crystallographers who refuse to believe that the element hydrogen even
exists. Never seen it. Including them in refinement has even been called
"too many parameters". Nowadays, of course, we know having hydrogens
involved in the geometry is a good thing. This is especially true at
poor resolution.
The other important hydrogen keyword to know is:
MAKE HYDR A
This is the default. "A" means add all hydrogens, regardless of what is
in the input pdb file. Normally, these get deleted before output, so
you don't notice it. The other choices are:
MAKE HYDR N: which is basically "do not do the hydrogen thing". Any
hydrogens in the input file will be ignored, and they will also not be
in the output file (even if you use MAKE HOUT Y), because they were
never there.
MAKE HYDR Y : this is the one that will use the hydrogens listed in the
pdb file. Here there lies a danger! Not every program names hydrogens
the same way, so some might be quietly ignored. The only way to know if
they were is to use MAKE HOUT Y.
So, if you want to define and use your own hydrogens through many rounds
of refmac, you want to use:
MAKE HYDR Y
MAKE HOUT Y
But, if your starting PDB has no hydrogens in it you want your first
round to have:
MAKE HYDR A
MAKE HOUT Y
If you inspect the occupancies of the output hydrogens you may find that
some are zero, and every once ina while one is somewhere between 1 and
0. I have not seen documentation for this, but it appears that refmac
might be quietly doing some sort of occupancy refinement on hydrogens
that may or may not be there. Titratable ones for example. The
philisophy explained to me once by Garib is that since it is twice as
bad to put in an electron that isn't there vs leaving out one that is
you want to err on the side of dropping hydrogens that don't seem to fit.
HTH,
-James Holton
MAD Scientist
On 5/20/2020 7:07 PM, Bernhard Rupp wrote:
Hi Fellows,
for an experiment, I am running 0.9 A data with unrestrained Refmac
(yes I know should/could use SHELXL, but let’s drop that for now).
When I select ‘ignore even if present in file’ in a PDB that has
hydrogens, I get the identical results than with ’use if present’ or
‘generate all’.
The log informs me that the instructions were properly issued, the
output PDB does not have Hs, but Rs and map are exactly the same
as with selection of if present or generate all H. Does not seem to
make sense.
If I cull the hydrogens from the input PDB and ‘use if present’ or
‘ignore’, the stats and map are different and no H in output as
requested - all as expected.
Maybe that can be reproduced and, if it is not a feature, fixed.
Best, BR
------------------------------------------------------
Bernhard Rupp
http://www.hofkristallamt.org/
b...@hofkristallamt.org <mailto:b...@hofkristallamt.org>
+1 925 209 7429
+43 676 571 0536
------------------------------------------------------
Real knowledge is to know
the extent of one's ignorance
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