Baich A 1971 The biosynthesis of proline in .

Hayzer DJ, Leisinger T 1980 The gene-enzyme relationships of proline biosynthesis in .

Hayzer DJ, Leisinger T 1981 Proline biosynthesis in .

AB - Radioisotopic assays were developed for the enzymes of proline metabolism. These assays are specific and sensitive enough to measure enzyme activities in cultured cells and biopsy specimens. Measurements of these enzymes in tissues and cultured cells suggest that endogenous biosynthesis of proline may be an important source of this amino acid.

Hayzer DJ, Leisinger T 1982 Proline biosynthesis in .

Hayzer DJ, Moses V 1978b The enzymes of proline biosynthesis in .

Uridylyltransferase is activated by -ketoglutarateand ATP, while it is inhibited by glutamine and Pi.The following diagram summarizes the regulation of bacterial glutaminesynthetase (see text page 1035) : We can "walk through" this regulatory cascade by looking at aspecific example, namely increased levels of -ketoglutarate( reflecting a corresponding increase in NH3) levels: (1) Uridylyltransferase activity is increased (2) PII (in complex with adenylyltransferase)is uridylylated (3) Glutamine synthetase is deadenylylated (4) -ketoglutarate and NH3 form glutamine and Pi That the control of bacterial glutamine synthetase is exquisitely sensitiveto the level of the cell's nitrogen metabolites is illustrated by the fact thatthe glutamine just produced in the above cascade is now an inhibitor of furtherglutamine production. Proline, Ornithine and Arginine are derived from GlutamateThe first step involves phosphorylation of glutamate by ATP with the enzyme -glutamylkinase, followed by reduction to glutamate-5-semialdehyde which spontaneouslycyclizes (no enzyme required) to an internal Schiff base. The formation ofthe semialdehyde also requires the presence of either NADP or NADPH.The semialdehyde is a branch point, however.

Uridylyltransferase is activated by -ketoglutarateand ATP, while it is inhibited by glutamine and Pi.The following diagram summarizes the regulation of bacterial glutaminesynthetase (see text page 1035) : We can "walk through" this regulatory cascade by looking at aspecific example, namely increased levels of -ketoglutarate( reflecting a corresponding increase in NH3) levels: (1) Uridylyltransferase activity is increased (2) PII (in complex with adenylyltransferase)is uridylylated (3) Glutamine synthetase is deadenylylated (4) -ketoglutarate and NH3 form glutamine and Pi That the control of bacterial glutamine synthetase is exquisitely sensitiveto the level of the cell's nitrogen metabolites is illustrated by the fact thatthe glutamine just produced in the above cascade is now an inhibitor of furtherglutamine production. Proline, Ornithine and Arginine are derived from GlutamateThe first step involves phosphorylation of glutamate by ATP with the enzyme -glutamylkinase, followed by reduction to glutamate-5-semialdehyde which spontaneouslycyclizes (no enzyme required) to an internal Schiff base. The formation ofthe semialdehyde also requires the presence of either NADP or NADPH.The semialdehyde is a branch point, however.


Saccharomyces cerevisiae proline biosynthesis

In summary, the biogenesis of proline is regulated by three PYCRs that have distinct sub-cellular localization and enzymatic properties. PYCR1 and PYCR2 are localized in the mitochondria, and primarily involved in the conversion of glutamate to proline, and are subject to product inhibition. PYCRL is a cytoplasmic enzyme, exclusively involved in conversion or ornithine to proline. This enzyme is not inhibited by proline. Now that we have established the role of each PYCR in proline biosynthesis, and have illuminated distinctions in their enzymatic properties, it will be possible to probe their role in melanoma and other diseases in an informed manner.

Ammonia Assimilation and Proline | Biosynthesis | Proline

This study also illustrates an important technical point on the silencing of metabolic enzymes with siRNA. In the case of these targets, one cannot always expect the knockdown with siRNA to produce a proportional inhibition of 13C enrichment from a substrate to a product. Indeed, the efficiency of gene knockdown by siRNA is usually 85–95%, so in almost all cases some enzyme is expressed. In fact, others have also observed less than complete inhibition of isotopic enrichment of a product from a substrate when silencing metabolic enzymes with shRNA [2,32]. If the enzyme in question is not the rate-limiting step, then even 5-10% remaining enzyme could support substantial conversion of a substrate into a product. This is likely to be part of the reason that knockdown of P5CS reduces the isotopic enrichment in proline from glutamate by nearly 90%, but knockdown of PYCRs fails to reach this degree. Furthermore, in this instance, where two PYCRs function along a particular path, knockdown of one isozyme is expected to only partially reduce the 13C enrichment in proline from the respective precursor.

Protein involved in the biosynthesis of the cyclic amino acid proline

One implication of this study, which is illustrated in the working model, is that P5C exists in separate unmixed pools. Although the steady-state level of P5C is too low to be measured directly, the inference of separate P5C pools follows from the results of gene silencing experiments. Thus, knockdown of PYCRL reduces the isotopic enrichment in proline from ornithine, but not from glutamate. However, if P5C, which is the common intermediate along both routes, existed as a single pool, then knockdown of PYCRL would alter the isotopic enrichment in proline in the same way from both precursors. This observation is consistent with the idea that the observed P5C pool separation may reflect its channeling within multi-enzyme complexes that contain the PYCRs, as has been observed in other systems [9,23,24]. The fact that PYCRL functions exclusively in the cytoplasm to convert P5C to proline, raises questions about how P5C is generated in the cytosol. This remains an open issue because OAT is in the mitochondrial matrix, but we cannot exclude the possibility that some form of OAT exists in the cytosol and is coupled to PYCRL.