On 20 Aug 2008, at 18:24, James Pauff wrote:
> Hello all, thank you for the responses. Just to clear up a couple
> of things...
> 1) My dataset was acquired on a synchrotron and scaled/truncated
> there. To my knowledge they used the same procedure as for other
> structures that we have obtained there...which have had no B factor
> issues. Granted, our modified protein is of a different spacegroup
> than previous.
The first thing that I do with my data after coming back from the
synchrotron is process them nice and quite.
I know the selling point of synchrotrons that you go away with your
data processed all too well,
but the current reality of sample changers and ultra fast detectors
does not allow time to do the job on site
in an optimal fashion, in many cases. I would not be exaggerating to
say that every single dataset at NKI over the last 8 years,
we have processed again back at home to be sure we did things the
best possible way.
>
> 2) I have not used/touched the TLS parameters at all.
>
> The idea that our 73% completeness (thus lacking 27% of the
> 'weakest' reflections) has lead to an artificially low B factor
> sounds most appealing at this point?
Most likely the following is clear to you and most readers, but just
to be sure we are talking about the same things:
1. I was talking about Wilson B in my first answer, which would be
affected directly from missing weak reflections.
2. I see now you are talking about the mean B of all atoms, which I
am not sure how it would be affected by missing reflections
3. In your last email you go back (?) talking about Wilson B
(TRUNCATE) which is a whole different think than the mean B value of
all atoms you mentioned first.
Wilson B: property of your data, gradient of the fall-off of
Intensity as a function of resolution
Mean B of all atoms: property of your model, average B of all atoms
"For good data and a good model the two should not be too far apart"
Best, Tassos
> As usual, I greatly appreciate all of your insights here!
>
> Best,
> Jim
>
>
> --- On Wed, 8/20/08, Eleanor Dodson <[log in to unmask]> wrote:
>
>> From: Eleanor Dodson <[log in to unmask]>
>> Subject: Re: [ccp4bb] Lower completeness, decent R factors, but
>> low B factor...
>> To: [log in to unmask]
>> Cc: [log in to unmask]
>> Date: Wednesday, August 20, 2008, 4:30 AM
>> James Pauff wrote:
>>> Hello all,
>>>
>>> I have a refined structure at 2.6 angstroms that at
>> about 73% completeness at this resolution. The I/sigma is
>> about 2.0 at 2.6 angstroms, and the omit density for my
>> ligands is great contoured at 3.0sigma. My Rcryst is 19 or
>> so and the Rfree is 24.5 or so.
>>>
>>> HOWEVER, my mean B value is 13.9, whereas my other 2
>> structures (at 2.2 and 2.3 angstroms, same protein, >95%
>> completeness) have mean B values of 22+. Any suggestions as
>> to what is going on here? I'm having trouble explaining
>> this.
>>>
>>> Thank you,
>>> Jim
>>>
>>>
>>>
>>>
>>>
>>>
>> Have you used TLS - listed B factors will then be given
>> relative to the
>> TLS parameters. You need to run tLSANL to get a more
>> realistic value.
>> Eleanor
>>
>>
>> But in fact temperature factors are rather harder to
>> estimate at lower
>> resolutions than higher. Look at your <Fo> and
>> <Fc> curves v resolution
>> ( part of a REFMAC loggraph) and you can see that sometimes
>> the overall
>> scaling struggles to get a reasonable fit..
>
>
>
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