Structure of a TFIIB-TBP-TATA-element and its Assembly

by Jolee J. Passanante

Introduction:

Eukaryotic transcription, while sharing similarities with the analogous process in Prokaryotes, is notably more complex and as a result is less well characterized. Out of all eukaryotic transcription the best understood at the molecular level is for genes which are transcribed by RNA pol II. This process proceeds through the recognition of the TATA box region of the promoter by TFIID with the help of TBP. This forms the pre-initiation complex (PIC). TFIIB then enters the complex creating heterotrimeric protein/DNA structure that can then be recognized by a pol II complex. The paper in review has solved the crystal structure (2.7 A res) of human-TFIIB bound to TBP2 from Arabidopsis, which is in turn selectively binding to the adenovirus Major Late Promoter (AdMLP). Although in vivo the PIC begins with the TBP mediated positioning of TFIID in vitro studies show that it is possible to initiate transcription with only the TBP-TFIIB complex. This indicates that TBP and TFIIB can position Pol II alone. It is this TBP-TFIIB Pol II positioning complex which has been imaged here.

Assembly of the Complex:

1.

The first element needed for complex formation is the normal cellular B form DNA. Specifically, we are concerned with the TATA box region of the Adenovirus Major Late Promoter, as this is the region which is recognized by the TBP-TFIIB complex. The full sequence used in the crystallization is GGCTATAAAAGGGCTG (region of recognition is highlighted in yellow).

(if using Internet Explorer Press twice) Upon binding of the TBP, the DNA is distorted out of its native B conformation. The DNA becomes partly unwound and shows a lateral displacement of 18 angstroms and a 110 deg. angle between the axes of incoming and out going DNA. This large DNA deformation permits Pol II transcription factors to have closer associations than would be possible with linear DNA.

2.

This brings us to the TBP, the next addition to the complex. TBP is a monomeric saddle shaped protein composed of 2 almost structurally identical domains. Each of these domains is composed of 2 alpha helices and 5 stretches of beta sheet. It is this beta sheet region of the protein which interacts with the TATA box

and thus gives it its sequence specificity to type II promoters within the genome. The distortion results mainly from the intercalation of four Phenylalanine residues. There is one located on each of the "stirrups" and one on the beta turn linker region between the beta stretch 2 and 3.

The primary protein/DNA interaction is through hydrogen bonding patterns on the protein's surface that specifically match the h-bonding pattern in the minor grove of the TATA box region (successive pairs of H-bond acceptors). There are also interactions that are non-specific which are between the protein and the DNA backbone.

The third protein to enter the complex is the TFIIB. It is also a monomeric protein with two identical domains connected in tandem. Each domain consists of 5 alpha helices. The C terminal domain contains a short 6th helix on its carboxyl end.

TFIIB binds to the DNA which is bent since it is already in complex with the TBP.

4.The Full Structure

of the complex is now shown which leads us to the next section of the tour.

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Protein/DNA Interactions:

1. TBP/DNA:

We have already seen how TPBís four phenylalanine residues intercalate with DNA thus unwinding and bending it, but this explains little about itís specificity to the TATA box.

Upon closer examination it can be seen that the

binding is composed of hydrophobic interactions and H-bonding to both the backbone, in a non-sequence specific manner, and to the bases themselves in a very sequence specific manner.

2. TFIIB/DNA:

The interaction between the TFIIB protein and the TATA region is significantly less than that of TBP.

Not only are the TFIIB interactions more limited than those of the TBP but they are completely base non-specific because it only contacts the DNA backbone

as can now be seen when only amino acids which interact with the bases are shown,....there are none! This begs the question "How then is TFIIB binding specific to the TATA promoter region?" which brings us to the next set of interesting interactions.

Protein/Protein Interactions:

1.

The addition of TFIIB to the transcription complex is only specific to the TATA region because it binds specifically to TBP,as shown above, is sequence specific to the TATA region.

In particular the Protein/Protein recognition happens primarily between the intervening beta turn region of residues on H1', S2í and S3í of the carboxy end domain of TBP and regions on helices H3, H3í, H4, H5, and the interdomain linker region.

Which can be seen better in this view as the structure is represented in strands.

Recruitment of Other Transcription Factors

This is of course only the beginning! Once the complex has formed it can itself promote transcription by Pol II or can be affected by other transcription factors or even repressors. These other factors bind specifically to regions of the complex based on interactions such as charge affinity, H-bond patterning, van der Waals packing and of course surface shape recognition.

This last view of the structure shows the charge on the surface of the complex suggesting sites for interaction of these additional transcription factors. Red=neg. Blue=pos.

References:

D.B.Nikolov et al., Crystal structure of a TFIIB-TBP-TATA-element ternary complex; Nature, vol. 377 1995, 119-128.