setting solvent density threshold at 2.5 times of the bulk solvent density

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Mohan Pradhan

8 Sep 2015 8 Sep '15

1:52 a.m.

Dear Chimera users, I am using the Volume Viewer application in Chimera to visualize the solvent density around protein at a threshold value of 2.5 times of the bulk solvent density. My understanding is that the iso value corresponding to the bulk solvent density is marked by the peak in the plot of the volume viewer. I am then multiplying the iso value of the peak by 2.5Is this the right way of identifying regions in protein that have a solvent density of 2.5 times of the bulk solvent density? Kindly suggest. Thanks,Mohan

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Elaine Meng

8 Sep 8 Sep

9:14 a.m.

New subject: setting solvent density threshold at 2.5 times of the bulk solvent density

Dear Mohan, This sounds like a general issue with EM, independent of Chimera, and may depend on the specific data set, experimental conditions, resolution, etc. Unfortunately I don’t have an answer… perhaps some users who are expert in EM may be able to respond. Best, Elaine ---------- Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

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On Sep 8, 2015, at 1:52 AM, Mohan Pradhan wrote:

Dear Chimera users, I am using the Volume Viewer application in Chimera to visualize the solvent density around protein at a threshold value of 2.5 times of the bulk solvent density.

My understanding is that the iso value corresponding to the bulk solvent density is marked by the peak in the plot of the volume viewer. I am then multiplying the iso value of the peak by 2.5 Is this the right way of identifying regions in protein that have a solvent density of 2.5 times of the bulk solvent density?

Kindly suggest. Thanks, Mohan

Tom Goddard

1:07 p.m.

New subject: setting solvent density threshold at 2.5 times of the bulk solvent density

Hi Mohan, What you suggest is not correct. I don't know if your map is from x-ray crystallography or electron microscopy but in both cases the map values are not literally "density" (ie mass per unit volume). The processing of the maps often introduces negative values, and often puts the most common map value (peak histogram) at zero or even a negative value. As Elaine, suggested you would need to ask someone expert in the processing of maps. Chimera only uses the numeric values in the map file, it knows nothing about how those values relate to physical properties. I have never seen an attempt to quantify mass density with x-ray or EM of biological samples. X-ray researchers instead usually look at a iso-contour 1 or 1.5 or 2 standard deviations above the mean map value. Tom

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On Sep 8, 2015, at 1:52 AM, Mohan Pradhan wrote:

Dear Chimera users,

I am using the Volume Viewer application in Chimera to visualize the solvent density around protein at a threshold value of 2.5 times of the bulk solvent density.

My understanding is that the iso value corresponding to the bulk solvent density is marked by the peak in the plot of the volume viewer. I am then multiplying the iso value of the peak by 2.5 Is this the right way of identifying regions in protein that have a solvent density of 2.5 times of the bulk solvent density?

Kindly suggest.

Thanks, Mohan _______________________________________________ Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

Mohan Pradhan

10 Sep 10 Sep

9:20 a.m.

Dear Tom and Elaine, Thank you both for your response. I think I was not very clear with my question. Let me rephrase my query here again in more detail. I have done Molcular Dynamics simulation of my protein in explicit solvent. Following which I calculated the water density from the trajectory using grid-based approach with grid cells of size 0.5 X 0.5 X 0.5 Å. A sample of the grid-based density output file is attached here for your reference. We then used Chimera to visualize the map using the volume viewer menu. We are attaching two image files here at two different isocountor threshold cutoff. One which was set to the isovalue at which we observe the maximum value of the historgram at an isovalue of 37. (image-1) Second we set it to 2.5 times that isovalue 2.5 X 37 ~ 93) (Image-2). The issue we wanted to clarify was what do these histogram actually depict with regard to the input grid file? What is the histogram depicting at an iso-value of zero? Why is there a sinusodial curve for the histogram towards the lower isovlaue? Thanks,Mohan On Wednesday, September 9, 2015 4:07 AM, Tom Goddard wrote: Hi Mohan, What you suggest is not correct. I don't know if your map is from x-ray crystallography or electron microscopy but in both cases the map values are not literally "density" (ie mass per unit volume). The processing of the maps often introduces negative values, and often puts the most common map value (peak histogram) at zero or even a negative value. As Elaine, suggested you would need to ask someone expert in the processing of maps. Chimera only uses the numeric values in the map file, it knows nothing about how those values relate to physical properties. I have never seen an attempt to quantify mass density with x-ray or EM of biological samples. X-ray researchers instead usually look at a iso-contour 1 or 1.5 or 2 standard deviations above the mean map value. Tom On Sep 8, 2015, at 1:52 AM, Mohan Pradhan wrote: Dear Chimera users, I am using the Volume Viewer application in Chimera to visualize the solvent density around protein at a threshold value of 2.5 times of the bulk solvent density. My understanding is that the iso value corresponding to the bulk solvent density is marked by the peak in the plot of the volume viewer. I am then multiplying the iso value of the peak by 2.5Is this the right way of identifying regions in protein that have a solvent density of 2.5 times of the bulk solvent density? Kindly suggest. Thanks,Mohan_______________________________________________ Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

Elaine Meng

9:50 a.m.

New subject: setting solvent density threshold at 2.5 times of the bulk solvent density

Dear Mohan, Yes, that is a very different question now that we know you calculated the map from MD. Everything depends on how you calculated the “density" to produce the input map. Rather than an actual density, it may just be counts of how many times there was a water inside each grid cell (such as if you used the occupancy analysis in the MD Movie tool). If so, you could divide those values by the number of MD frames that the counts came from, to make them more meaningful; for example, a count of 40 for a sample of 100 MD frames means that a grid cell had a water in it 40% of the time. MD Movie occupancy analysis: http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/movie/movie.html#o... Volume Viewer is just displaying this raw data, it isn’t doing anything to transform or normalize it. The data is just a grid of points with associated values. The histogram shows the range of values horizontally, and the bar heights show how many grid points have those values. It is not useful to show the contour surface for values near zero, and I would not read too much into the shape of the histogram. The high value near zero just means many grid points have values of zero, and the other maximum just means many grid points have values of 37. If you use a different size grid cell and/or sampled a different number of MD frames (if the values are simply counts), the maximum would be a different value. I hope this helps, Elaine ---------- Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

...

On Sep 10, 2015, at 9:20 AM, Mohan Pradhan wrote:

Dear Tom and Elaine, Thank you both for your response. I think I was not very clear with my question. Let me rephrase my query here again in more detail.

I have done Molcular Dynamics simulation of my protein in explicit solvent. Following which I calculated the water density from the trajectory using grid-based approach with grid cells of size 0.5 X 0.5 X 0.5 Å. A sample of the grid-based density output file is attached here for your reference.

We then used Chimera to visualize the map using the volume viewer menu. We are attaching two image files here at two different isocountor threshold cutoff. One which was set to the isovalue at which we observe the maximum value of the historgram at an isovalue of 37. (image-1)

Second we set it to 2.5 times that isovalue 2.5 X 37 ~ 93) (Image-2).

The issue we wanted to clarify was what do these histogram actually depict with regard to the input grid file?

What is the histogram depicting at an iso-value of zero?

Why is there a sinusodial curve for the histogram towards the lower isovlaue?

Thanks, Mohan

Tom Goddard

1:01 p.m.

New subject: setting solvent density threshold at 2.5 times of the bulk solvent density

Hi Mohan, I think your occupancy map histogram has a peak at 0 and peak at 37 is explained as follows. The peak near means there are many grid points in the volume that water never reaches. This must be the volume occupied by the protein. The peak at 37 are grid points outside the protein where usually 37 waters are found per grid point over the course of your MD frames. So I think you are correctly interpreting the values. Using a threshold 2.5 * 37 would show grid points where waters are found 2.5 times more often than average — you might think of those locations as “bound waters” — I’d expect them to be all at points adjacent to the protein. One extra bit of info about the histogram, the horizontal axis is the occupancy value (0-354 for your data) and the vertical axis is the number of grid points having a specific occupancy value. The horizontal axis is linear, but the vertical axis is logarithmically scaled. So on the vertical axis a slightly higher peak represents many times more grid points. Tom

...

On Sep 10, 2015, at 9:50 AM, Elaine Meng wrote:

Dear Mohan, Yes, that is a very different question now that we know you calculated the map from MD.

Everything depends on how you calculated the “density" to produce the input map. Rather than an actual density, it may just be counts of how many times there was a water inside each grid cell (such as if you used the occupancy analysis in the MD Movie tool). If so, you could divide those values by the number of MD frames that the counts came from, to make them more meaningful; for example, a count of 40 for a sample of 100 MD frames means that a grid cell had a water in it 40% of the time. MD Movie occupancy analysis: http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/movie/movie.html#o...

Volume Viewer is just displaying this raw data, it isn’t doing anything to transform or normalize it. The data is just a grid of points with associated values. The histogram shows the range of values horizontally, and the bar heights show how many grid points have those values. It is not useful to show the contour surface for values near zero, and I would not read too much into the shape of the histogram. The high value near zero just means many grid points have values of zero, and the other maximum just means many grid points have values of 37. If you use a different size grid cell and/or sampled a different number of MD frames (if the values are simply counts), the maximum would be a different value.

I hope this helps, Elaine ---------- Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

...
On Sep 10, 2015, at 9:20 AM, Mohan Pradhan wrote:

Dear Tom and Elaine, Thank you both for your response. I think I was not very clear with my question. Let me rephrase my query here again in more detail.

I have done Molcular Dynamics simulation of my protein in explicit solvent. Following which I calculated the water density from the trajectory using grid-based approach with grid cells of size 0.5 X 0.5 X 0.5 Å. A sample of the grid-based density output file is attached here for your reference.

We then used Chimera to visualize the map using the volume viewer menu. We are attaching two image files here at two different isocountor threshold cutoff. One which was set to the isovalue at which we observe the maximum value of the historgram at an isovalue of 37. (image-1)

Second we set it to 2.5 times that isovalue 2.5 X 37 ~ 93) (Image-2).

The issue we wanted to clarify was what do these histogram actually depict with regard to the input grid file?

What is the histogram depicting at an iso-value of zero?

Why is there a sinusodial curve for the histogram towards the lower isovlaue?

Thanks, Mohan

_______________________________________________ Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

Mohan Pradhan

10:48 p.m.

Dear Tom and Elaine, Thank you for your response. It helped me a lot in clarifying my concepts about volume viewer and solvent density maps. Best Regards,Mohan On Friday, September 11, 2015 4:01 AM, Tom Goddard wrote: Hi Mohan, I think your occupancy map histogram has a peak at 0 and peak at 37 is explained as follows. The peak near means there are many grid points in the volume that water never reaches. This must be the volume occupied by the protein. The peak at 37 are grid points outside the protein where usually 37 waters are found per grid point over the course of your MD frames. So I think you are correctly interpreting the values. Using a threshold 2.5 * 37 would show grid points where waters are found 2.5 times more often than average — you might think of those locations as “bound waters” — I’d expect them to be all at points adjacent to the protein. One extra bit of info about the histogram, the horizontal axis is the occupancy value (0-354 for your data) and the vertical axis is the number of grid points having a specific occupancy value. The horizontal axis is linear, but the vertical axis is logarithmically scaled. So on the vertical axis a slightly higher peak represents many times more grid points. Tom

...

On Sep 10, 2015, at 9:50 AM, Elaine Meng wrote:

Dear Mohan, Yes, that is a very different question now that we know you calculated the map from MD.

Everything depends on how you calculated the “density" to produce the input map. Rather than an actual density, it may just be counts of how many times there was a water inside each grid cell (such as if you used the occupancy analysis in the MD Movie tool). If so, you could divide those values by the number of MD frames that the counts came from, to make them more meaningful; for example, a count of 40 for a sample of 100 MD frames means that a grid cell had a water in it 40% of the time. MD Movie occupancy analysis: http://www.rbvi.ucsf.edu/chimera/docs/ContributedSoftware/movie/movie.html#o...

Volume Viewer is just displaying this raw data, it isn’t doing anything to transform or normalize it. The data is just a grid of points with associated values. The histogram shows the range of values horizontally, and the bar heights show how many grid points have those values. It is not useful to show the contour surface for values near zero, and I would not read too much into the shape of the histogram. The high value near zero just means many grid points have values of zero, and the other maximum just means many grid points have values of 37. If you use a different size grid cell and/or sampled a different number of MD frames (if the values are simply counts), the maximum would be a different value.

I hope this helps, Elaine ---------- Elaine C. Meng, Ph.D. UCSF Computer Graphics Lab (Chimera team) and Babbitt Lab Department of Pharmaceutical Chemistry University of California, San Francisco

...
On Sep 10, 2015, at 9:20 AM, Mohan Pradhan wrote:

Dear Tom and Elaine, Thank you both for your response. I think I was not very clear with my question. Let me rephrase my query here again in more detail.

I have done Molcular Dynamics simulation of my protein in explicit solvent. Following which I calculated the water density from the trajectory using grid-based approach with grid cells of size 0.5 X 0.5 X 0.5 Å. A sample of the grid-based density output file is attached here for your reference.

We then used Chimera to visualize the map using the volume viewer menu. We are attaching two image files here at two different isocountor threshold cutoff. One which was set to the isovalue at which we observe the maximum value of the historgram at an isovalue of 37. (image-1)

Second we set it to 2.5 times that isovalue 2.5 X 37 ~ 93) (Image-2).

The issue we wanted to clarify was what do these histogram actually depict with regard to the input grid file?

What is the histogram depicting at an iso-value of zero?

Why is there a sinusodial curve for the histogram towards the lower isovlaue?

Thanks, Mohan

_______________________________________________ Chimera-users mailing list Chimera-users@cgl.ucsf.edu http://plato.cgl.ucsf.edu/mailman/listinfo/chimera-users

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Elaine Meng
Mohan Pradhan
Tom Goddard