Repressor molecules

Modulation of activator and repressor function by effector molecules. Repressors can either be inhibited (A) or stimulated (B) by effector-molecule interactions. Similarly, activators can either be stimulated (C) or inhibited (D) by effector-molecule interactions. Transcription-factor functions can also be modulated by posttranslational modification such as phosphorylation (see Fig. 29.10). 

Folding of the polypeptide chain as it grows may require special small molecules, like appropriate coenzymes, for proper coiling. Small-molecule repressors may be molecules that cause improper folding of the growing polypeptide chain. 

In the above section we studied a model that could nicely explain how the expression of a gene can be controlled by modifying the number of regulatory molecules repressors. The problem with this model is that it relies upon an assumption that... 

Figure 7-15 shows the time evolution of the temperature, total energy, and potential energy for a 300 ps simulation of the tetracycline repressor dimer in its induced (i.e., hgand-bound) form. Starting from the X-ray structure of the monomer in a complex with one molecule of tetracycline and a magnesium ion (protein database... 

The fundamental unit of tertiary structure is the domain. A domain is defined as a polypeptide chain or a part of a polypeptide chain that can fold independently into a stable tertiary structure. Domains are also units of function. Often, the different domains of a protein are associated with different functions. For example, in the lambda repressor protein, discussed in Chapter 8, one domain at the N-terminus of the polypeptide chain binds DNA, while a second domain at the C-terminus contains a site necessary for the dimerization of two polypeptide chains to form the dimeric repressor molecule. 

This model of Cro binding to DNA was arrived at by intuition and clever model building. Its validity was considerably strengthened when the same features were subsequently found in the DNA-binding domains of the lambda-repressor molecule. The helix-turn-helix motif with a recognition helix is present in the repressor, and moreover the repressor DNA-binding domains dimerize in the crystals in such a way that the recognition helices are separated by 34 A as in Cro. 

The 434 Cro molecule contains 71 amino acid residues that show 48% sequence identity to the 69 residues that form the N-terminal DNA-binding domain of 434 repressor. It is not surprising, therefore, that their three-dimensional structures are very similar (Figure 8.11). The main difference lies in two extra amino acids at the N-terminus of the Cro molecule. These are not involved in the function of Cro. By choosing the 434 Cro and repressor molecules for his studies, Harrison eliminated the possibility that any gross structural difference of these two molecules can account for their different DNA-binding properties. 

Figure 8.19 The a helices of the N-terminal region of the trp repressor are involved in subunit interactions and form a stable core in the middle of the dimer. Alpha helices 4-6, which include the helix-turn-helix motif, form two "head" regions at the two ends of the molecule. Alpha helix 3 connects the core to the head in both subunits. (Adapted from R.W. Schevitz et al., Nature 317 782-786, 1985.)...

More than 30 years ago Jacob and Monod introduced the Escherichia coli lac operon as a model for gene regulation. The lac repressor molecule functions as a switch, regulated by inducer molecules, which controls the synthesis of enzymes necessary for E. coli to use lactose as an energy source. In the absence of lactose the repressor binds tightly to the operator DNA preventing the synthesis of these enzymes. Conversely when lactose is present, the repressor dissociates from the operator, allowing transcription of the operon. 

The tetrameric structure of the lac repressor has a quite unusual V-shape (Figure 8.22). Each arm of the V-shaped molecule is a tight dimer, which is very similar in structure to the PurR dimer and which has the two N-termi-nal DNA binding domains close together at the tip of the arm. The two dimers of the lac repressor are held together at the other end by the four carboxy-terminal a helices, which form a four-helix bundle. 

Figure 8.22 The lac repressor molecule is a V-shaped tetramer in which each arm is a dimer containing a DNA-hinding site. The helix-tum-helix motifs (red) of each dimer bind in two successive major grooves and the hinge helices (purple) bind adjacent to each other in the minor groove between the two major groove binding sites. The four subunits of the tetramer are held together by the four C-terminal helices (yellow) which form a four helix bundle. The bound DNA fragments are bent. (Adapted from M. Lewis et al., Science 271 1247-1254, 1996.)...

Some of the procaryotic DNA-binding proteins are activated by the binding of an allosteric effector molecule. This event changes the conformation of the dimeric protein, causing the helix-tum-helix motifs to move so that they are 34 A apart and able to bind to the major groove. The dimeric repressor for purine biosynthesis, PurR, induces a sharp bend in DNA upon binding caused by insertion of a helices in the minor groove between the two... 

Like Thr 124 and Thr 215, the Asn 69 and Asn 159 residues occupy equivalent positions in the two homologous motifs of TBP. By analogy with the symmetric binding of a dimeric repressor molecule to a palindromic sequence described in Chapter 8, the two motifs of TBP form symmetric sequence-specific hydrogen bonds to the quasi-palindromic DNA sequence at the center of the TATA box. The consensus TATA-box sequence has an A-T base pair at position 4, but either a T-A or an A-T base pair at the symmetry-related position 5, and the sequence is, therefore, not strictly palindromic. However, the hydrogen bonds in the minor groove can be formed equally well to an A-T base pair or to a T-A base pair, because 02 of thymine and N3 of adenine occupy nearly stereochemically equivalent positions, and it is sufficient, therefore, for the consensus sequence of the TATA box to be quasi-palindromic. 

Acquired resistance has been observed by constitutive upregulation of mdr efflux pump expression due to a mutation inactivating a respective repressor or inducibly, caused by molecules transiently inactivating repressor molecules upon binding. Depending upon the substrate spectra of the respective subset of efflux pumps upregulated, a multiple drug resistance (mdr) phenotype is expressed, which in combination with a specific resistance mechanism can contribute to a clinically relevant level of resistance. 

PIAS (protein inhibitors of activated STATs) proteins were first discovered in yeast-two-hybrid screens as interacting molecules with STAT transcription factors. The mammalian family consists ofthe founding member PIAS3, which was described as a repressor of STAT3, and three additional members, PIAS1, PIASy (also known as PIAS4), and PIASx (also known as... 

The product of the repressor gene, the 236-amino-acid, 27 IcDa repressor protein, exists as a two-domain molecule in which the amino terminal domain binds to operator DNA and the carboxyl terminal domain promotes the association of one repressor protein with another to form a dimer. A dimer of repressor molecules binds to operator DNA much more tighdy than does the monomeric form (Figure 39-6A to 39-6C). 

Detailed analysis of the lambda repressor led to the important concept that transcription regulatory proteins have several functional domains. For example, lambda repressor binds to DNA with high affinity. Repressor monomers form dimers, dimers interact with each other, and repressor interacts with RNA polymerase. The protein-DNA interface and the three protein-protein interfaces all involve separate and distinct domains of the repressor molecule. As will be noted below (see Figure 39—17), this is a characteristic shared by most (perhaps all) molecules that regulate transcription. 

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