User:Marianne Weulersse/Sandbox 202
From Proteopedia
Pauline TRISTRAM and Marianne WEULERSSE: THE HUMAN SRY-DNA COMPLEX
Contents |
Introduction
The human protein SRY (hSRY) is a transcription factor implicated in the differentiation of gonads during development. SRY is encoded by the SRY gene located on the human chromosome Y and consists of 223 residues which comprises three domains: N-terminal domain, central DNA-binding domain called High Mobility Group (HMG) box and the C-terminal domain.
The molecular structure of the human SRY-HMG-DNA complex was determined by double and triple multi-dimensional NMR spectroscopy.
|
Molecular structure
The general hSRY structure[1]
The molecular structure of the human SRY-HMG-DNA complex was determined by double and triple multi-dimensional NMR spectroscopy. The protein consists of 3 α-helical segments (, and ) which form an L-shaped structure stabilized by an . The of the ‘‘L’’consists of the extended N-terminal section and the helix 3 along with the C-terminal region which are oriented anti-parallel to each other. The N-terminal tail is anchored to the end of helix 3 and the beginning of the C-terminal tail by an . The consists of helixes 1 and 2.
The binding domain
The binds specifically DNA sequence through the minor groove (more hydrophobic than the major groove, especially the adenine base which has no -NH2 group). Its sequence specificity is related with included in the conserved basic terminal tail of the HMG box [2], which trigger the C-terminal region to bend toward the N-terminus and with packed together.
Protein binding and conformation changes
Structure modification of the complex
The DNA flexibility and hydrophobicity are two required key elements of the DNA sequence for the specific binding that occurs especially through the protein sidechains (more flexible than the backbone)[3]. ‘‘Induced Fit’’ conformational changes exist in the process of hSRY-HMG domain-DNA binding . Both hSRY and DNA change their conformations to achieve greater complementarity of geometries and properties during the binding process sequence specificity, thus promoting a perfect specific complex ( PDB entry 1hrz)
- In the free state , the extended N/C-terminus and the helix 3 of the HMG domain are very flexible, and the DNA have limited and uniform internal mobility.
- In the complex, the protein backbone is less mobile, whereas the DNA backbone is more mobile (the two ends moved slightly and bases A3 and T14 twisted).
The specific binding site occurs exclusively in the minor groove and triggers a large conformational change. The aliphatic sidechain of Ile13, like a hydrophobic wedge at the apex, partially intercalates between A5 and A6 bases and widens the minor groove (about 5Å). DNA becomes underwounded (70–80° bending) and its conformation gets gradually closer to A-DNA (narrowing of the major groove about 3Å).The original free B-DNA highly hydrated loses water molecules after the hydrophobic interaction with the HMG domain under the influence of its hydrophobic cores . More over, the A-like conformation of the DNA increases the hydrophobic contacts between protein and the DNA.
Interaction between DNA and hSRY-HMG : complex structure strengthening
Hydrophobic Recognition
The structural complementarity[4] of DNA with hSRY-HMG domain promotes specific interactions (hydrophobic interactions and salt bridges) that contribute to stabilize the bent DNA, the protein itself and the whole complex. Hydrophobic recognition is the physical basis of HMG–DNA interaction that cause primarily the DNA conformation change.The two hydrophobic cores of the hSRY-HMG domain maintained stable contacts with DNA through hydrophobic interactions with DNA backbone and bases.
- The is exposed on the concave surface at the apex of the L-shaped structure [5] . It consists of aromatic residues located at the junction of the three helices. The sidechains stabilize all three helices because of a strong and large delocalized Π interaction which form a stable edgeto-face interaction with the Π planes of the DNA A5 and A6 bases.
The major structural change occurs in the long arm of the L-shaped conformation of the hSRY-HMG domain. The angle between the long and short arms is more obtuse to accommodate the DNA through the sway of the extended N- and C-termini. The aromatic sidechains of Phe12, packed against DNA bases, lay at the horizontal level with which deform the DNA. Met9 and Trp43 bind the sugar phosphate backbone thereby opening the minor groove. is an important aliphatic residue of the big hydrophobic core located at the joint of the extended N-terminal and the helix 1, close to the intercalating residue Ile13 and in contact with the backbone of DNA. Ile13 could be affected by the sway of Met9 and becomes more feasible to intercalate into the basepairs of DNA. Met9 mutations would result in a decrease of the binding affinity and/or bending angles. The naturally occuring mutation Met9 => Ile would reduce the swing because of the shorter sidechain of Ile( -CH3 branch in the Cα atom). The swing of Ile is therefore too weak to help the intercalation of Ile13. As a result, the angle of DNA bending decreases and the affinity of DNA binding reduces. The conformation change (deviations) of the short loop region around Met9 is essential to the function of SRY.
- The at the C-terminal and N-terminal regions consists of Val5, Tyr69, Tyr72, and Tyr74. These residues link the two termini together and are important for the sequence-specific binding to DNA.
Tyr74 interacts with the base of A3 and pushes the base toward the major groove.
Electrostatics interactions
The surrounding hydrophobic region consists of positively charged residues which form electrostatic interactions (salt bridges) with the phosphate groups of DNA. Such favors the deformation of DNA and the formation of a stable complex. Many carboxyamide groups of SRY-HMG residues are also involved in electrostatic interactions with nitrogen atoms of DNA bases (Asn10 interacts with the N2 atom of G13), thereby triggering the conformation change and the stability of the complex.
Hydrogen bonds
The hydroxyl groups of residues are used to develop water-mediated hydrogen bonds with the nitrogen atoms of DNA bases (Ser33 forms a hydrogen bond with the N2 atom of G9).They participate to the specific SRY-HMG-DNA interaction and allow the strong bond of the protein.
- There are five direct hydrogen bonds (one between the hydroxyl group of Ser36 and the O2 atom of T10 and the others between the protein and DNA phosphate backbone).
- One is located between Asn10 and the O2 atom of C4 and T14.
Sequence conservation
Many sequencing showed that the SRY protein differs from one species to another except for the HMG domain that remains preserved.The main influence of the SRY gene regulation comes from the structural alteration of the promoter of the target DNA.
Conclusion
SRY acts as a transcriptional activator by binding specifically to the sequence DNA octamer target site AACAA-(A/T)(G/C) of the MIS gene promoter (Müllerian Inhibiting Substance). Therefore, the conformational change of the DNA latter allows it to be recognized by other transcription factors inducing the transcription of genes implicated in the development of the male reproductive system. The MIS gene codes for a substance which cause regression of the female genital tract such as the precursor of the uterus, upper vagina and fallopian tube. SRY may act also as a transcriptional repressor by binding the cytochrome P450 aromatase promoter, enabling the conversion of testosterone to estradiol in order to promote the development of the testes.
References
- ↑ Werner MH, Huth JR, Gronenborn AM, Clore GM. Molecular basis of human 46X,Y sex reversal revealed from the three-dimensional solution structure of the human SRY-DNA complex. Cell. 1995 Jun 2;81(5):705-14. PMID:7774012
- ↑ Phillips NB, Jancso-Radek A, Ittah V, Singh R, Chan G, Haas E, Weiss MA. SRY and human sex determination: the basic tail of the HMG box functions as a kinetic clamp to augment DNA bending. J Mol Biol. 2006 Apr 21;358(1):172-92. Epub 2006 Feb 6. PMID:16504207 doi:10.1016/j.jmb.2006.01.060
- ↑ Murphy EC, Zhurkin VB, Louis JM, Cornilescu G, Clore GM. Structural basis for SRY-dependent 46-X,Y sex reversal: modulation of DNA bending by a naturally occurring point mutation. J Mol Biol. 2001 Sep 21;312(3):481-99. PMID:11563911 doi:http://dx.doi.org/10.1006/jmbi.2001.4977
- ↑ Bianchi ME, Beltrame M. Flexing DNA: HMG-box proteins and their partners. Am J Hum Genet. 1998 Dec;63(6):1573-7. PMID:9837808 doi:10.1086/302170
- ↑ Tang Y, Nilsson L. Interaction of human SRY protein with DNA: a molecular dynamics study. Proteins. 1998 Jun 1;31(4):417-33. PMID:9626701
