A second issue we and other groups have encountered is the potential for LSD1 inhibitors to inhibit HRP. 1 This peptide is a potent inhibitor of LSD1, however, this methodology was not available at the time our manuscript was published. It is now known that the LSD1 reaction can be quenched using the 21-mer peptide in which dimethyllysine 4 is replaced by a methionine. The first issue, which is manageable, is that there is no reagent in the kit with which to quench the enzymatic reaction, and thus the timing of each step in the procedure is critical to obtaining reliable results. The radical cation is then quantitated with Amplex Red. Recombinant LSD1 and the peptide substrate are co-incubated, and the activity of the enzyme is determined by converting the LSD1 by-product H 2O 2 to a radical cation using horseradish peroxidase (HRP). The assay employs a 21-mer peptide substrate analogous to the tail found on histone 3, containing dimethylated lysine at position 4. The LSD1 assay procedures in the manuscript were conducted using a commercially available LSD1 assay kit (BPS Bioscience, San Diego, CA, kit #50106). Since the original publication of this manuscript, we have encountered problems with the assay procedure that was used to gather the data for the kinetic analysis of compound 6d. The correct graph is shown in the figure, generated as described below. However, the data obtained in the absence of inhibitor was inadvertently omitted from the figure. 4 on page 1605, a Lineweaver-Burk graph was inserted that indicated compound 6d competitively inhibited lysine-specific demethylase 1 (LSD1) with a Ki of 2.4 μM. The table below summarizes the possibilities.The Authors regret that in Fig. The other oxygens in the backbone can only act as hydrogen bond acceptors. The oxygens at 7 and 12 can act as either donors or acceptors. Unlike the backbone of a protein, RNA backbone doesn't contain any nitrogen atoms, so hydrogen bonding must involve oxygen atoms. In DNA, the oxygen at atom 12 is not present, and the first base atom is 12. The first atom of the base in RNA is atom 13. Atom numbering continues normally from there, and no "virtual atoms" are involved. Since there's no next RNA segment, the oxygen at atom 9 is not attached to a phosphate group. Foldit still identifies atoms 1 through 4 as being present, and these "virtual atoms" can even be banded.įor the last RNA segment in a chain, numbering is more straightforward. The first atom visible is atom 5, a carbon attached to the ribose ring. Atom 12 is the oxygen in the hydroxyl group, which Foldit considers to be the first "sidechain" atom, meaning it's not part of the backbone.įor the first RNA segment in a chain, the phosphate group is missing. Atom 10 is a carbon (carbon 1'), which is where the actual RNA base is attached. Atom 9 is an oxygen which joins the phosphate group of the next RNA segment. Foldit considers one of the oxygen atoms of the phosphate to belong to the ribose of the preceding segment, so only the oxygen atoms 2 through 4 are numbered.Ītom numbering continues with the ribose group. Foldit starts numbering with the phosphorous in the phosphate group. For example, the C terminal of a chain has an extra backbone atom, so it's beta carbon, the first atom of the sidechain, is 6 instead of 5.įor RNA, the backbone atoms always have the same number, regardless position in the chain. In a protein, the number of atoms in a segment is different depending on its position in the chain. The rules for numbering atoms are different in RNA, however. Just as in a protein, Foldit can identify atom numbers in RNA. The hydroxyl group is what distinguishes RNA from DNA. A hydroxyl group (oxygen and hydrogen) is attached to carbon 2'. The RNA base is attached to carbon 1' of the ribose. The carbon atoms are referred to as 1', 2', 3' and 4'. The sugar group in the RNA backbone is always a form of ribose, a pentagonal ring consisting of an oxygen atom and four carbon atoms. Each segment of RNA or DNA is linked to the next by a phosphate. A phosphate group consists of a phosphorous atom bound to four oxygen atoms. This is why "RNA" stands for "ribonucleic acid", and "DNA" stands for "deoxyribonucleic acid".īoth RNA and DNA backbone consists of alternating groups phosphates and sugars. The difference is that RNA backbone contains the sugar ribose, while DNA backbone contains the sugar deoxyribose. RNA backbone is highly similar to DNA backbone. The backbone of RNA is more complicated than the backbone of the amino acids in a protein. The blue numbers 1' to 4' refer to the carbons in the in ribose ring. [So u know this is not true ,oxygen can only make two bonds so it (oxygen)could not be top of pyramid in a tetrahedron formation.only carbon can make four bonds[Ifįile:Rna_|thumb|400px| RNA backbone showing chemical groups.
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