Photograph 51: How a blurry X revealed the structure of DNA

 So those of you who read my previous article may have looked at the conveniently included Photograph 51 and wondered how on that shows a double helix.

For those who have not read my previous article, here is another image of it! 

                                       

It may look blurry and vague- but this is one of the most important photos ever produced in the history of genomics- possibly even in the history of science in general!

This photo was produced by Rosalind Franklin and Raymond Gosling, using a technique called X-ray Crystallography. What this, in very simple terms, means is firing X-rays at something and seeing how they bounce back at you. Franklin and Gosling carried this out on DNA fibres and because this was the 51st photograph they had created, they named it photograph 51. They had to expose the DNA to X-rays for 62 hours. For reasons we will NOT go into (mainly because I did it last week and I am still exhausted), James Watson and Francis Crick got hold of this photograph and created their model of DNA. 

This is what the double helix of DNA looks like:

 

This image also very usefully includes the four nitrogenous bases that make up the 'rungs'. Each colour corresponds to the base in question and only an Adenine can bind to a Thymine, whilst a Cytosine can only bind to the Guanine. Meanwhile, the 'ladder' is made up of phosphate; what this image does not include is the sugar. Between the phosphate and the nitrogenous bases, we have a type of sugar called Deoxyribose. That's actually how we get the full name of DNA: Deoxyribonucleic acid.

 

When Watson and Crick published, they used the following image:

 

  

 

So, the question is: How does that X in photograph 51 correspond to this double stranded helix we now know and love at the structure of DNA? Well, the image is rather complicated and there is a lot going on. But stay tuned and hopefully it will make a bit more sense! 

So, the first thing to cover is two terms: Diffraction and Interference. 

Diffraction is what is used to describe the scattering of X-rays when they hit the molecule in question. So, x-rays applied to DNA fibres would scatter back. Interference is when these scattered X-rays combine with each other. This can be both 'constructive' or 'destructive'. When its constructive interference, the x-rays combine in such a way that it creates a stronger wave. This results in a stronger signal - which we see as bright spots on the image. When its destructive interference, the X-rays cancel each other out - and that means you don't get any signal at all. You are just left with black spots. In X-ray crystallography, you will have a pattern of constructive and destructive interference - in other words, a pattern of black spots and bright spots. This gives clues on the structure.

 

So, looking at photograph 51 again:

 

 

The black dots that make an X, along with the black smudges at the top and bottom (yes, they are part of the image!) are examples of destructive interference, whilst the lighter patches are constructive. You will see that the spots of destructive interference are regularly spaced and repeating. There's the same number of black spots either side and they are all equally spaced. If you were to look at a double helix from the side, you would see it looking somewhat like a zigzag.

Something like this:

So, X-rays are fired at this zigzag and ping back off. Because the spaces of the zigzag are regularly spaced apart, this means the X-rays combine constructively and destructively at regular intervals. The x-rays ping off and combine at right angles to the helix; this happens at regular intervals, giving us that cross we see in photograph 51. 

 

 

The image above I got from Brian Sutton writing for King's College London on April 2023 and may show it better than I can. What we see in black on the blue background is the wavy zigzag a helix from the side would present as. The red lines are the x-rays pinging off the helix. Some ping in an upwards direction whilst some goes downwards. Where they meet is marked by a little red X. These are the areas of destructive interference- which we see as black spots on Photograph 51. 

So now we know that we must have a helical structure. But there is still a bit more information we can get from the image! I did say this image was so important!

If you were to measure the vertical space between the spots and then measure the distance between the centre of the image to either the top or bottom 'smudge', you would find that the vertical space is 1/10 of the distance between the centre of the image to the top or bottom smudge. These smudges are caused by base stacking in the middle of the helix; this means the bases are arranging themselves, so their electrons are overlapping and increasing the strength of their interactions. Based on the distances, it was concluded that each turn of the helix must contain 10 stacked bases. Franklin said that it was 'highly probable' that the bases were located inside the helix- remember, that was not even known then! She also predicted that the phosphate bases must therefore be located on the outside of the helix. 

But it is still not the end of the story!! Looking at this photo, Watson and Crick realised that base stacking was happening in DNA and so they began to wonder how the bases could arrange themselves for this to happen. This led them to the concept of base pairing; Adenine can only go to Thymine, and Cytosine can only go to Guanine. I do have to admit that as far as I know, Rosalind Franklin had not quite thought of base-pairing, so I do (rather grudgingly) must give credit for that one to Watson and Crick. But they would not have done it without her image!! 

Are we at the end of what we can deduce with Photograph 51? Of course not! Rosalind Franklin, whilst Watson and Crick were building their model, noticed that there is a space in each of the arms of the cross where a fourth spot should be. Based on this, she predicted that the two chains would be separated by three-eighths’ of the pitch of the double helix. 

This is where we finally come to an end on what Photograph 51 can tell us. Hopefully, this blog post has given some insight into why it was so important and why it was so outrageous that Franklin never got the proper credit for the photograph. Watson and Crick were only able to do their model because of this photograph!