subject:"\[ccp4bb\] Extra density close to phosphate bound to Zn2\+"

Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-06 Thread Bernhard Rupp

One practically HAS to do what Dale describes. The sigma levels of a difference 
map peak depends on how much it is above the mean noise level of the map. With 
high resolution (and given a cooperating molecule) it is very likely that the 
model is very good and complete, thus low mean map noise level, and anything 
missing will create huge peaks. A forgotten water molecule then can easily have 
difference peak (sigma not absolute) levels in the 20s... 

Best, BR

-Original Message-
From: CCP4 bulletin board  On Behalf Of Dale Tronrud
Sent: Tuesday, August 6, 2019 00:24
To: CCP4BB@JISCMAIL.AC.UK
Subject: Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

   One thing to remember is that Li+ only has two electrons.  It should be a 
little more than twice the density of an hydrogen (because the charge pulls in 
the electron cloud).  If you are seeing lithium at 8 "sigma" you should be 
seeing the well located hydrogen atoms at 4 "sigma".  Are you?

   I always like controls.  Take out one of the near by water molecules and see 
how that, known, difference peak compares to your mystery peak.
 A Li+ should have about 1/5th the height of an H2O.  If you like the 
hypothesis of two orientations of the PO4 group, the relative height of the two 
peaks will give insight to the occupancy ratio.

   Remember, if the PO4 has two orientations in the crystal, the water 
molecules it (they?) is/are bound to will likely also have alternatives.

Dale Tronrud

On 8/5/2019 6:05 AM, Maria Håkansson wrote:
> Dear CCP4 bulletin board,
> 
> I am working with some lytic enzymes called endolysines, which bind 
> Zn2+ in the active site. I have three homologues protein determined to 
> 1.2 Å each where the Zn2+ is bound to a cystein, two histidines and 
> one phosphate ion added (1.9-2.3 Å binding distances) in the 
> crystallization experiment.
> 
> Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 
> sigma peak is present in all three endolysines, see below.
> I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 
> 13Å2) ion.
> Sodium has benn added in the crystallization experiments since sodium 
> potassium phosphate salt has been used. The only reason for including 
> Li+ is that I think the binding distances (1.7-2.0 Å) are too short 
> for Na+.
> 
> I have also tried to make a model with the phosphate in two different 
> conformations but it does not fit.
> 
> Have anyone seen something similar before? What is the most correct 
> way of dealing with unknown densities?
> It is difficult to disregard +8 sigma difference density close to the 
> active site.
> 
> Thanks in advance for any help!
> 
> Best regards,
> Maria
> 
>  
> 
> Maria Håkansson, PhD, Crystallization Facility Manager Principal 
> Scientist
> 
> SARomics Biostructures AB
> Medicon Village
> SE-223 81 Lund, Sweden
> 
> Mobile: +46 (0)76 8585706
> Web: www.saromics.com <http://www.saromics.com>
> 
> 
> 
> 
> 
> --
> --
> 
> To unsubscribe from the CCP4BB list, click the following link:
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> 

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Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-05 Thread Dale Tronrud

   One thing to remember is that Li+ only has two electrons.  It should
be a little more than twice the density of an hydrogen (because the
charge pulls in the electron cloud).  If you are seeing lithium at 8
"sigma" you should be seeing the well located hydrogen atoms at 4
"sigma".  Are you?

   I always like controls.  Take out one of the near by water molecules
and see how that, known, difference peak compares to your mystery peak.
 A Li+ should have about 1/5th the height of an H2O.  If you like the
hypothesis of two orientations of the PO4 group, the relative height of
the two peaks will give insight to the occupancy ratio.

   Remember, if the PO4 has two orientations in the crystal, the water
molecules it (they?) is/are bound to will likely also have alternatives.

Dale Tronrud

On 8/5/2019 6:05 AM, Maria Håkansson wrote:
> Dear CCP4 bulletin board,
> 
> I am working with some lytic enzymes called endolysines, which
> bind Zn2+ in the active site. I have three homologues protein
> determined to 1.2 Å each where the Zn2+ is bound
> to a cystein, two histidines and one phosphate ion added (1.9-2.3 Å
> binding distances) in the crystallization experiment.
> 
> Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 sigma
> peak is present in all three endolysines, see below.
> I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 13Å2)
> ion. 
> Sodium has benn added in the crystallization experiments since sodium
> potassium phosphate
> salt has been used. The only reason for including Li+ is that I think
> the binding distances (1.7-2.0 Å) are too short for Na+.
> 
> I have also tried to make a model with the phosphate in two different
> conformations but it does not fit.
> 
> Have anyone seen something similar before? What is the most correct way
> of dealing with unknown densities?
> It is difficult to disregard +8 sigma difference density close to the
> active site.
> 
> Thanks in advance for any help!
> 
> Best regards,
> Maria
> 
>  
> 
> Maria Håkansson, PhD, Crystallization Facility Manager
> Principal Scientist
> 
> SARomics Biostructures AB
> Medicon Village
> SE-223 81 Lund, Sweden
> 
> Mobile: +46 (0)76 8585706
> Web: www.saromics.com 
> 
> 
> 
> 
> 
> 
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
> 

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[ccp4bb] Fwd: [ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-05 Thread Daniel M. Himmel, Ph. D.

-- Forwarded message -
From: Daniel M. Himmel, Ph. D. 
Date: Mon, Aug 5, 2019 at 12:45 PM
Subject: Re: [ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to
phosphate bound to Zn2+
To: 


Pentagonal phosphate geometry would be a transition state, except that
three oxygens
would be more or less co-planar.  Would anything approaching a transition
state show up in the electron
density of a crystal structure?  In myosin ATPase structures, there are
phosphate analogs with geometry
that approximates a transition state.

-Daniel


On Mon, Aug 5, 2019 at 11:55 AM  wrote:

> Hi Maria,
>
>
>
> Did you rotate the phosphate, or invert it? If you invert the phosphate,
> you may get into trouble with the parameters. Although a phosphate is
> symmetric, its oxygens have different names and inverting it leads to all
> kind of problems, especially in a high resolution map which does not allow
> the phosphate to flip it back in an “allowed” conformation.
>
>
>
> If you did rotate the phosphate, I take my words back and you may have to
> look for ions.
>
>
>
> Best,
>
> Herman
>
>
>
> *Von:* CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] *Im Auftrag von
> *Maria Håkansson
> *Gesendet:* Montag, 5. August 2019 17:32
> *An:* CCP4BB@JISCMAIL.AC.UK
> *Betreff:* [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound
> to Zn2+
>
>
>
> *EXTERNAL : *Real sender is owner-ccp...@jiscmail.ac.uk
>
>
>
> Hi,
>
>
>
> Thanks for your suggestions but I have tried to invert the phosphate and
> it is not
>
> fitting the map. The geometry is not correct that way and it is too good
> data to ignore and
>
> to my knowledge a pentacoordinated phosphate is a non existent species.
>
>
>
> That leaves me with ions.
>
>
>
> Best regards,
>
> Maria
>
>
>
>
>
> Maria Håkansson, PhD, Crystallization Facility Manager
> Principal Scientist
>
> SARomics Biostructures AB
> Medicon Village
> SE-223 81 Lund, Sweden
>
> Mobile: +46 (0)76 8585706
> Web: www.saromics.com
> <https://urldefense.proofpoint.com/v2/url?u=http-3A__www.saromics.com=DwMFaQ=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ=n8AawGvnjHilw4Yoh7AjCCk8wfameakvWM7OvpbvCJI=q-G0klB2rvoNDw6bKWKr8ASbOLmfrjWlRry1Hsms-54=>
>
>
>
>
>
>
>
>
>
> On 5 Aug 2019, at 16:32, Jan Abendroth  wrote:
>
>
>
> Hi Maria,
>
> apart from the suggestions that were already made, take a look at the P-O
> bond on the right side of the P. Maybe it is just the perspective, this
> appears to be rather long.
>
> So, instead of the alternate conformation, maybe just a flip of the
> phosphate and a water off to the right?
>
>
>
> Cheers,
>
> Jan
>
>
>
> On Mon, Aug 5, 2019 at 7:16 AM Nukri Sanishvili 
> wrote:
>
> Hi Maria,
>
> Let's not ignore the "missing" density - the red one. If the question is
> only P vs S, a sulfur would add to the negative density, i.e. make matters
> worse. It also appears that modeling a phosphate in alternative
> conformations, as suggested by Wim and Roger, would take care of the issue
> of negative density as well.
>
> Cheers,
>
> Nukri
>
>
>
> On Mon, Aug 5, 2019 at 8:05 AM Maria Håkansson <
> maria.hakans...@saromics.com> wrote:
>
> Dear CCP4 bulletin board,
>
>
>
> I am working with some lytic enzymes called endolysines, which
>
> bind Zn2+ in the active site. I have three homologues protein
>
> determined to 1.2 Å each where the Zn2+ is bound
>
> to a cystein, two histidines and one phosphate ion added (1.9-2.3 Å
> binding distances) in the crystallization experiment.
>
>
>
> Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 sigma
> peak is present in all three endolysines, see below.
>
> I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 13Å2)
> ion.
>
> Sodium has benn added in the crystallization experiments since sodium
> potassium phosphate
>
> salt has been used. The only reason for including Li+ is that I think the
> binding distances (1.7-2.0 Å) are too short for Na+.
>
>
>
> I have also tried to make a model with the phosphate in two different
> conformations but it does not fit.
>
>
>
> Have anyone seen something similar before? What is the most correct way of
> dealing with unknown densities?
>
> It is difficult to disregard +8 sigma difference density close to the
> active site.
>
>
>
> Thanks in advance for any help!
>
>
>
> Best regards,
>
> Maria
>
>
>
>   ion.png>
>
>
>
> Maria Håkansson, PhD, Crystallization Facility M

[ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-05 Thread Herman . Schreuder

PS, you also have to define an alternative conformation, but that you probably
already did.

Von: Schreuder, Herman /DE
Gesendet: Montag, 5. August 2019 17:55
An: 'Maria Håkansson'; CCP4BB@JISCMAIL.AC.UK
Betreff: AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to
Zn2+

Hi Maria,

Did you rotate the phosphate, or invert it? If you invert the phosphate, you
may get into trouble with the parameters. Although a phosphate is symmetric,
its oxygens have different names and inverting it leads to all kind of
problems, especially in a high resolution map which does not allow the
phosphate to flip it back in an “allowed” conformation.

If you did rotate the phosphate, I take my words back and you may have to look
for ions.

Best,
Herman

Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Maria
Håkansson
Gesendet: Montag, 5. August 2019 17:32
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

EXTERNAL : Real sender is owner-ccp...@jiscmail.ac.uk

Hi,

Thanks for your suggestions but I have tried to invert the phosphate and it is
not
fitting the map. The geometry is not correct that way and it is too good data
to ignore and
to my knowledge a pentacoordinated phosphate is a non existent species.

That leaves me with ions.

Best regards,
Maria

Maria Håkansson, PhD, Crystallization Facility Manager
Principal Scientist

SARomics Biostructures AB
Medicon Village
SE-223 81 Lund, Sweden

Mobile: +46 (0)76 8585706
Web:
www.saromics.com<https://urldefense.proofpoint.com/v2/url?u=http-3A__www.saromics.com=DwMFaQ=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ=n8AawGvnjHilw4Yoh7AjCCk8wfameakvWM7OvpbvCJI=q-G0klB2rvoNDw6bKWKr8ASbOLmfrjWlRry1Hsms-54=>

On 5 Aug 2019, at 16:32, Jan Abendroth
mailto:jan.abendr...@gmail.com>> wrote:

Hi Maria,
apart from the suggestions that were already made, take a look at the P-O bond
on the right side of the P. Maybe it is just the perspective, this appears to
be rather long.
So, instead of the alternate conformation, maybe just a flip of the phosphate
and a water off to the right?

Cheers,
Jan

On Mon, Aug 5, 2019 at 7:16 AM Nukri Sanishvili
mailto:sannu...@gmail.com>> wrote:
Hi Maria,
Let's not ignore the "missing" density - the red one. If the question is only P
vs S, a sulfur would add to the negative density, i.e. make matters worse. It
also appears that modeling a phosphate in alternative conformations, as
suggested by Wim and Roger, would take care of the issue of negative density as
well.
Cheers,
Nukri

On Mon, Aug 5, 2019 at 8:05 AM Maria Håkansson
mailto:maria.hakans...@saromics.com>> wrote:
Dear CCP4 bulletin board,

I am working with some lytic enzymes called endolysines, which
bind Zn2+ in the active site. I have three homologues protein
determined to 1.2 Å each where the Zn2+ is bound
to a cystein, two histidines and one phosphate ion added (1.9-2.3 Å binding
distances) in the crystallization experiment.

Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 sigma peak
is present in all three endolysines, see below.
I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 13Å2) ion.
Sodium has benn added in the crystallization experiments since sodium potassium
phosphate
salt has been used. The only reason for including Li+ is that I think the
binding distances (1.7-2.0 Å) are too short for Na+.

I have also tried to make a model with the phosphate in two different
conformations but it does not fit.

Have anyone seen something similar before? What is the most correct way of
dealing with unknown densities?
It is difficult to disregard +8 sigma difference density close to the active
site.

Thanks in advance for any help!

Best regards,
Maria

Maria Håkansson, PhD, Crystallization Facility Manager
Principal Scientist

SARomics Biostructures AB
Medicon Village
SE-223 81 Lund, Sweden

Mobile: +46 (0)76 8585706
Web:
www.saromics.com<https://urldefense.proofpoint.com/v2/url?u=http-3A__www.saromics.com_=DwMFaQ=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ=n8AawGvnjHilw4Yoh7AjCCk8wfameakvWM7OvpbvCJI=iDeElvjjAtCzkGvlyWvkljE9fucoGgiM-zXxXE2PnL0=>

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[ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-05 Thread Herman . Schreuder

Hi Maria,

Did you rotate the phosphate, or invert it? If you invert the phosphate, you 
may get into trouble with the parameters. Although a phosphate is symmetric, 
its oxygens have different names and inverting it leads to all kind of 
problems, especially in a high resolution map which does not allow the 
phosphate to flip it back in an “allowed” conformation.

If you did rotate the phosphate, I take my words back and you may have to look 
for ions.

Best,
Herman

Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Maria 
Håkansson
Gesendet: Montag, 5. August 2019 17:32
An: CCP4BB@JISCMAIL.AC.UK
Betreff: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+


EXTERNAL : Real sender is owner-ccp...@jiscmail.ac.uk

Hi,

Thanks for your suggestions but I have tried to invert the phosphate and it is 
not
fitting the map. The geometry is not correct that way and it is too good data 
to ignore and
to my knowledge a pentacoordinated phosphate is a non existent species.

That leaves me with ions.

Best regards,
Maria


Maria Håkansson, PhD, Crystallization Facility Manager
Principal Scientist

SARomics Biostructures AB
Medicon Village
SE-223 81 Lund, Sweden

Mobile: +46 (0)76 8585706
Web: 
www.saromics.com<https://urldefense.proofpoint.com/v2/url?u=http-3A__www.saromics.com=DwMFaQ=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ=n8AawGvnjHilw4Yoh7AjCCk8wfameakvWM7OvpbvCJI=q-G0klB2rvoNDw6bKWKr8ASbOLmfrjWlRry1Hsms-54=>





On 5 Aug 2019, at 16:32, Jan Abendroth 
mailto:jan.abendr...@gmail.com>> wrote:

Hi Maria,
apart from the suggestions that were already made, take a look at the P-O bond 
on the right side of the P. Maybe it is just the perspective, this appears to 
be rather long.
So, instead of the alternate conformation, maybe just a flip of the phosphate 
and a water off to the right?

Cheers,
Jan

On Mon, Aug 5, 2019 at 7:16 AM Nukri Sanishvili 
mailto:sannu...@gmail.com>> wrote:
Hi Maria,
Let's not ignore the "missing" density - the red one. If the question is only P 
vs S, a sulfur would add to the negative density, i.e. make matters worse. It 
also appears that modeling a phosphate in alternative conformations, as 
suggested by Wim and Roger, would take care of the issue of negative density as 
well.
Cheers,
Nukri

On Mon, Aug 5, 2019 at 8:05 AM Maria Håkansson 
mailto:maria.hakans...@saromics.com>> wrote:
Dear CCP4 bulletin board,

I am working with some lytic enzymes called endolysines, which
bind Zn2+ in the active site. I have three homologues protein
determined to 1.2 Å each where the Zn2+ is bound
to a cystein, two histidines and one phosphate ion added (1.9-2.3 Å binding 
distances) in the crystallization experiment.

Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 sigma peak 
is present in all three endolysines, see below.
I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 13Å2) ion.
Sodium has benn added in the crystallization experiments since sodium potassium 
phosphate
salt has been used. The only reason for including Li+ is that I think the 
binding distances (1.7-2.0 Å) are too short for Na+.

I have also tried to make a model with the phosphate in two different 
conformations but it does not fit.

Have anyone seen something similar before? What is the most correct way of 
dealing with unknown densities?
It is difficult to disregard +8 sigma difference density close to the active 
site.

Thanks in advance for any help!

Best regards,
Maria

 

Maria Håkansson, PhD, Crystallization Facility Manager
Principal Scientist

SARomics Biostructures AB
Medicon Village
SE-223 81 Lund, Sweden

Mobile: +46 (0)76 8585706
Web: 
www.saromics.com<https://urldefense.proofpoint.com/v2/url?u=http-3A__www.saromics.com_=DwMFaQ=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ=n8AawGvnjHilw4Yoh7AjCCk8wfameakvWM7OvpbvCJI=iDeElvjjAtCzkGvlyWvkljE9fucoGgiM-zXxXE2PnL0=>






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--
Jan Abendroth
Emerald Bio

Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-05 Thread Eleanor Dodson

I always say this, but is there any feature in the anomalous map? You can
sometimes see S atoms.

Eleanor

And do the temperatre factors suggest anything about multiple occupancies
etc..
E

On Mon, 5 Aug 2019 at 16:32, Maria Håkansson 
wrote:

> Hi,
>
> Thanks for your suggestions but I have tried to invert the phosphate and
> it is not
> fitting the map. The geometry is not correct that way and it is too good
> data to ignore and
> to my knowledge a pentacoordinated phosphate is a non existent species.
>
> That leaves me with ions.
>
> Best regards,
> Maria
>
>
> Maria Håkansson, PhD, Crystallization Facility Manager
> Principal Scientist
>
> SARomics Biostructures AB
> Medicon Village
> SE-223 81 Lund, Sweden
>
> Mobile: +46 (0)76 8585706
> Web: www.saromics.com
>
>
>
>
> On 5 Aug 2019, at 16:32, Jan Abendroth  wrote:
>
> Hi Maria,
> apart from the suggestions that were already made, take a look at the P-O
> bond on the right side of the P. Maybe it is just the perspective, this
> appears to be rather long.
> So, instead of the alternate conformation, maybe just a flip of the
> phosphate and a water off to the right?
>
> Cheers,
> Jan
>
> On Mon, Aug 5, 2019 at 7:16 AM Nukri Sanishvili 
> wrote:
>
>> Hi Maria,
>> Let's not ignore the "missing" density - the red one. If the question is
>> only P vs S, a sulfur would add to the negative density, i.e. make matters
>> worse. It also appears that modeling a phosphate in alternative
>> conformations, as suggested by Wim and Roger, would take care of the issue
>> of negative density as well.
>> Cheers,
>> Nukri
>>
>> On Mon, Aug 5, 2019 at 8:05 AM Maria Håkansson <
>> maria.hakans...@saromics.com> wrote:
>>
>>> Dear CCP4 bulletin board,
>>>
>>> I am working with some lytic enzymes called endolysines, which
>>> bind Zn2+ in the active site. I have three homologues protein
>>> determined to 1.2 Å each where the Zn2+ is bound
>>> to a cystein, two histidines and one phosphate ion added (1.9-2.3 Å
>>> binding distances) in the crystallization experiment.
>>>
>>> Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 sigma
>>> peak is present in all three endolysines, see below.
>>> I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 13Å2)
>>> ion.
>>> Sodium has benn added in the crystallization experiments since sodium
>>> potassium phosphate
>>> salt has been used. The only reason for including Li+ is that I think
>>> the binding distances (1.7-2.0 Å) are too short for Na+.
>>>
>>> I have also tried to make a model with the phosphate in two different
>>> conformations but it does not fit.
>>>
>>> Have anyone seen something similar before? What is the most correct way
>>> of dealing with unknown densities?
>>> It is difficult to disregard +8 sigma difference density close to the
>>> active site.
>>>
>>> Thanks in advance for any help!
>>>
>>> Best regards,
>>> Maria
>>>
>>>  >> ion.png>
>>>
>>> Maria Håkansson, PhD, Crystallization Facility Manager
>>> Principal Scientist
>>>
>>> SARomics Biostructures AB
>>> Medicon Village
>>> SE-223 81 Lund, Sweden
>>>
>>> Mobile: +46 (0)76 8585706
>>> Web: www.saromics.com
>>>
>>>
>>>
>>>
>>>
>>> --
>>>
>>> To unsubscribe from the CCP4BB list, click the following link:
>>> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>>>
>>
>> --
>>
>> To unsubscribe from the CCP4BB list, click the following link:
>> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
>>
>
>
> --
> Jan Abendroth
> Emerald Biostructures
> Seattle / Bainbridge Island, WA, USA
> home: Jan.Abendroth_at_gmail.com 
> work: JAbendroth_at_embios.com 
> http://www.emeraldbiostructures.com
>
>
>
> --
>
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
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Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

2019-08-05 Thread Maria Håkansson

Hi,

Thanks for your suggestions but I have tried to invert the phosphate and it is 
not
fitting the map. The geometry is not correct that way and it is too good data 
to ignore and
to my knowledge a pentacoordinated phosphate is a non existent species.

That leaves me with ions.

Best regards,
Maria


Maria Håkansson, PhD, Crystallization Facility Manager
Principal Scientist

SARomics Biostructures AB
Medicon Village
SE-223 81 Lund, Sweden

Mobile: +46 (0)76 8585706
Web: www.saromics.com




> On 5 Aug 2019, at 16:32, Jan Abendroth  wrote:
> 
> Hi Maria,
> apart from the suggestions that were already made, take a look at the P-O 
> bond on the right side of the P. Maybe it is just the perspective, this 
> appears to be rather long.
> So, instead of the alternate conformation, maybe just a flip of the phosphate 
> and a water off to the right?
> 
> Cheers,
> Jan
> 
> On Mon, Aug 5, 2019 at 7:16 AM Nukri Sanishvili  > wrote:
> Hi Maria,
> Let's not ignore the "missing" density - the red one. If the question is only 
> P vs S, a sulfur would add to the negative density, i.e. make matters worse. 
> It also appears that modeling a phosphate in alternative conformations, as 
> suggested by Wim and Roger, would take care of the issue of negative density 
> as well.
> Cheers,
> Nukri
> 
> On Mon, Aug 5, 2019 at 8:05 AM Maria Håkansson  > wrote:
> Dear CCP4 bulletin board,
> 
> I am working with some lytic enzymes called endolysines, which
> bind Zn2+ in the active site. I have three homologues protein
> determined to 1.2 Å each where the Zn2+ is bound
> to a cystein, two histidines and one phosphate ion added (1.9-2.3 Å binding 
> distances) in the crystallization experiment.
> 
> Now to my question. Close to the phosphate (B-factor=20Å2) ion a 8 sigma peak 
> is present in all three endolysines, see below.
> I have modeled it to a Na+ (B-factor= 30 Å2) or a Li+ (B factor = 13Å2) ion. 
> Sodium has benn added in the crystallization experiments since sodium 
> potassium phosphate
> salt has been used. The only reason for including Li+ is that I think the 
> binding distances (1.7-2.0 Å) are too short for Na+.
> 
> I have also tried to make a model with the phosphate in two different 
> conformations but it does not fit.
> 
> Have anyone seen something similar before? What is the most correct way of 
> dealing with unknown densities?
> It is difficult to disregard +8 sigma difference density close to the active 
> site.
> 
> Thanks in advance for any help!
> 
> Best regards,
> Maria
> 
>   ion.png>
> 
> Maria Håkansson, PhD, Crystallization Facility Manager
> Principal Scientist
> 
> SARomics Biostructures AB
> Medicon Village
> SE-223 81 Lund, Sweden
> 
> Mobile: +46 (0)76 8585706
> Web: www.saromics.com 
> 
> 
> 
> 
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 
> 
> To unsubscribe from the CCP4BB list, click the following link:
> https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 
> 
> 
> -- 
> Jan Abendroth
> Emerald Biostructures
> Seattle / Bainbridge Island, WA, USA
> home: Jan.Abendroth_at_gmail.com 
> work: JAbendroth_at_embios.com
> http://www.emeraldbiostructures.com 




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Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

[ccp4bb] Fwd: [ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

[ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

[ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

Re: [ccp4bb] Extra density close to phosphate bound to Zn2+

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