Fig. Structure of P. fluorescens phox. (A) Carbon representation of P. fluoresce
ID: 74266 • Letter: F
Question
Fig. Structure of P. fluorescens phox. (A) Carbon representation of P. fluorescens PhoX viewed from above the active site (left) or from the side (left) or from the side (right). The blades of the beta propeller are shown in different colours and two alpa subdomains are colored ray. The active-site ions are represented by green (Ca^2+). magenta Fe^3+, and cay (O^2_) spheres. (B)Clipped surface representation of Phox active site containing bound phosphate. Interatomic separations that are within bondin distance are shown between the matal ions and protein side chains (gray dashed lines and between the phosphate ion and coordinating groups (black dashed lines). These distances are tabulated in table s3. The structure of Pho X(papers 10 and 11) shows even greater differences vs. the PAPS:a Ca^2+ is added to an Fe^3+ -Fe^3+ center analogous to that in PAPs to ive a trianular trimetallic center; the tyrosinate liand to one of the Fe^3+ ions is replaced by a cysteinate ligand; and two Ca^2+ ions are located adjacent to the trinuclear center. What is the likely effect of the Tyr right arrow Cys substitution on the properties of the Fe^3+? Will the Cays ligand perform the same function you proposed for the Tyr in question l(b)? Is it reasonable that the CysS^- -fe^3+ unit in Pho X will result in the same color (purple) as the TyrO^- -Fe^3+ unit in PAP? What do you think happens to the enzyme when it is treated with EDTA (Fi. IC in paper 10), and why does subsequent addition of a solution of CaCl_2 restore the oriinal optical spectrum? The EPR spectrum of the native enzyme shows essentially no signal, consistent with the presence of two antiferromanetically coupled Fe^3+ ions, as in the PAPs. Addition of a powerful reductant (sodium dithionite), does not, however, result in the characteristic EPR signal around g=1.8 observed for the diiron PAPs. The authors suest that one Fe is released from the enzyme must contain an Fe^3+ center. Based on the authors" data and everything you have read about PAPs, do you find their argument convincing? Why or why not? If not, can you come up with an alternative explanation for their results?Explanation / Answer
A. Fe3+ ion is ligated by a tyrosine ligand results into intense phenolate to fe(III) charge transfer transition which is responsible for characteristic colour purple to the bacteria. So it there is substitution of the by cys then the Fe3+ will be unable to ligate to the amino acid as the binding of fe3+ ion to cys is unusual and phoX is the only protein that use the thiolate group coordinate a redox inactive fe3+ ion.and hence there will be no reaction involved which gives purple colour to bacteria.
Yes Cys will perform the same the same function as it also coordinates the fe3+ ion.
But there is no purple colour as in case of the Fe3+tyr ligand because additional thiolate to fe3+ charge transfer the bands from cysteine ligand will also contribute to the visible spectrum.
B. When enzyme is treated with EDTA,there is unfolding of the enzyme. And block the enzyme activity.so no density is seen corresponding to a molecule of EDTA in electron density maps.
Calcium ion helps in refolding of the PAPs enzyme as they are involved in the accessibility of the active site by the conformational change with a loop movement. So it leads to the restoration of the original spectrum and the density is visible in electron density maps.
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