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This question is related to something that happens when 3-phosphoglyceraldehyde is converted to 1,3-bisphosphoglycerate during glycolysis.

As I understand it, 3-phosphoglyceraldehyde is deprotonated by NAD+ and phosphorylated by a Pi group, which results in NADH + H. The extra H apparently comes from the Pi group (which I believe is called orthophosphate [1]).

Now, where does the Pi come from? is it just sitting in the cytosol?, does all Pi have a hydrogen? can someone clarify what an orthophosphate is, I never heard of that term.

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    $\begingroup$ You can google orthophosphoric acid. Pi is the acronym of inorganic phosphate. $\endgroup$ – Roni Saiba Oct 9 '17 at 3:33
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Yes, the cytosol contains a high concentration of orthophosphate (in yeast, between 10--75 mM) so it is readily available for this reaction. The notation P$_i$ is commonly used because the orthophosphate ion can exist in multiple forms in water solution:

H$_3$PO$_4$ $\leftrightarrow$ H$_2$PO$_4^{-}$ + H$^+$ $\leftrightarrow$ HPO$_4^{2-}$ + 2H$^+$$\leftrightarrow$ PO$_4^{3-}$ + 3H$^+$

When we don't need to specify an exact form, P$_i$ is convenient as a shorthand for "some phosphate ion".

At the cytosolic pH (about 7.2), almost all orthophosphate is in the H$_2$PO$_4^{-}$ and HPO$_4^{2-}$ forms, which are roughly equally abundant. So yes, there is (at least) one hydrogen on orthophosphate available for the enzymatic reaction.

The term "orthophosphate" rather than just "phosphate" is common in biochemistry because phosphate also exists as polymers, including the di- and triphophates (in nucleotides, for example) as well as longer polyphosphate chains. These form are all referred to as "phosphates", while "orthophophate" specifically means the "monomer" P$_i$.

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