Maybe somehow do partial cys partial cme, refine occupancies—is this possible in refmac?

JPK

From: CCP4 bulletin board [mailto:[log in to unmask]] On Behalf Of sreetama das
Sent: Saturday, January 17, 2015 3:11 PM
To: [log in to unmask]
Subject: Re: [ccp4bb] additional density on cysteine residue

Hi,
Thanks for the quick replies.

I modelled in CME and refined. However, I get a blob of negative density around the S-S bond, which is retained up to 5 sigma.

Does it mean it is not CME ?

Thanks & regards,

sreetama

On Sunday, 18 January 2015 12:41 AM, Roger Rowlett <[log in to unmask]> wrote:

If CSO does not account for the density -- the SO bond should be about 1.8 A IIRC -- a possibility is an adventitious metal ion.

Roger Rowlett

On Jan 17, 2015 1:25 PM, "sreetama das" <[log in to unmask]> wrote:

Dear Users,

I am solving a structure from x-ray diffraction data (1.62A resolution).

The protein has a single cysteine residue (which is also the catalytic residue), and it has a positive density on it (fig 1; R/Rfree = 16.88/19.94). The positive density is retained upto 11.5 sigma level.

Modelling with water retains the positive density (fig 2; R/Rfree = 16.85/19.94) upto 5.2 sigma level.

Modelling with CSO (S-hydroxycysteine, fig 3, R/Rfree = 16.82/ 19.81) produces partial positive and negative densities, which are retained upto 5 sigma. Moreover, after real-space refinement in coot followed by refinement in refmac, the N-terminus of CSO is not bonded to the preceding residue, nor is its C-terminus bonded to the succedding residue.

All maps are contoured at 1sigma (2Fo-Fc map) and 3sigma (fo-fc map).

The protein preparation contained Tris buffer at pH 7.2, NaCl, glycerol and beta-mercaptoethanol (2mM), while the crystallization condition contained citric acid (pH 3.5) and ammonium sulfate.

Please suggest how to interpret the data.

thanking in advance,

sreetama