Carbamoyl phosphate synthetase I

carbamoyl-phosphate synthetase 1, mitochondrial
Identifiers
Symbol CPS1
Entrez 1373
HUGO 2323
OMIM 608307
RefSeq NM_001875
UniProt P31327
Other data
EC number 6.3.4.16
Locus Chr. 2 p

Carbamoyl Phosphate Synthetase I is a ligase enzyme located in the mitochondria involved in the production of urea. Carbamoyl phosphate synthetase I (CPS1 or CPSI) transfers an ammonia molecule from glutamine or glutamate to a molecule of bicarbonate that has been phosphorylated by a molecule of ATP. The resulting carbamate is then phosphorylated with another molecule of ATP. The resulting molecule of carbamoyl phosphate leaves the enzyme.

Structure of Carbamoyl Phosphate Synthetase I

In E. coli CPSI is a heterodimer with a small subunit and a larger subunit with about 382 and 1073 amino acid residues in size, although in mammals (and other vertebrates) the CPSI protein is encoded by a single gene.[1] The small subunit contains one active site for the binding and deamination of glutamine to make ammonia and glutamate. The large subunit contains two active sites, one for the production of carboxyphosphate, and the other for the production of carbamoyl phosphate.[2][3] Within the large subunit there are two domains (B and C) each with an active site of the ATP-grasp family.[1] Connecting the two subunits is a tunnel of sorts, which directs the ammonia from the small subunit to the large subunit.[4]


Mechanism of Carbamoyl Phosphate Synthetase I

The overall reaction that occurs in CPSI is:

2ATP + HCO3− + NH4+ --> 2ADP + Carbamoyl phosphate + Pi[4]

This reaction can be thought of occurring in four distinct steps.[5]

  1. Bicarbonate is phosphorylated
  2. Ammonia is cleaved from glutamine (glutaminase) or glutamate (glutamate dehydrogenase)
  3. The ammonia attacks the carboxyphosphate, resulting in carbamate
  4. Carbamate is phosphorylated to give Carbamoyl phosphate

Of these four steps, only step two - the deamination of glutamine to get ammonia - is known to have actively participating amino acid residues, Cys269 and His353. The other three steps mostly utilize amino acid residue to form hydrogen bonds with substrates. A video of a simplified version of this mechanism is available here

Recent Mechanism Studies

It has been found that both ATP-binding sites in the large subunit of CPSI are structurally equivalent. A recent study has investigated the interlinking between these two domains (domain B and domain C) and has found evidence that they are coupled. This ATP-binding domain coupling works in a way such that a molecule of ATP binding at one site (domain C) conformationally allows synthesis at the other domain (domain B). If this is the case, carbamoyl phosphate is, in fact, not formed in step 5 (of the included mechanism below) by ejecting ADP but rather in step 4 by protonating the alcohol group and then kicking it off as water.[6]

Formation of carbamoyl phosphate mechanism
Deamination of glutamine to form ammonia and glutamate[7]

Regulation

CPSI is regulated by N-acetylglutamate; however, unlike most enzymes it is not allosterically regulated. Instead N-acetylglutamate is an obligate activator of CPS1.

Carbamoyl Phosphate Synthetase I and Metabolism

CPSI plays a vital role in protein and nitrogen metabolism. Once ammonia has been brought into the mitochondria via glutamine or glutamate, it is CPSI’s job to add the ammonia to bicarbonate along with a phosphate group to form carbamoyl phosphate. Carbamoyl phosphate is then put into the urea cycle to eventually create urea. Urea can then be transferred back to the blood stream and to the kidneys for filtration and on to the bladder for excretion.[8]

Health Problems related to Carbamoyl Phosphate Synthetase I

The main problem related to CPSI is genetics-based. Sometimes the body does not produce enough CPSI due to a mutation in the genetic code, resulting in poor metabolism of proteins and nitrogen, as well as high levels of ammonia in the body. This is dangerous because ammonia is highly toxic to the body, especially the nervous system, and can result in retardation and seizures.

References

  1. 1 2 James B. Thoden‡, Xinyi Hua, et al. “Carbamoyl-phosphate Synthetase: Creation of an Escape Route for Ammonia.” From the ‡Department of Biochemistry, University of Wisconsin, Madison, Wisconsin, 53706-1544 and the §Department of Chemistry, Texas A&M University, College Station, Texas 77843-3012. http://www.jbc.org/cgi/reprint/277/42/39722.pdf
  2. ↑ SUE GLENN POWERS, “Inhibition of Carbamyl Phosphate Synthetase by PI, P5-Di (adenosine 5’) - pentaphosphate EVIDENCE FOR TWO ATP BINDING SITES.” From the Department ofBiochemistry, Cornell University Medical College, New York, New York 10021. http://www.jbc.org/cgi/reprint/252/10/3558
  3. ↑ Thoden, James B.; Holden, Hazel M.; Wesenberg, Gary; Raushel, Frank M.; Rayment, Ivan (May 1997). "Structure of Carbamoyl Phosphate Synthetase: A Journey of 96 Å from Substrate to Product". Biochemistry. 36 (21): 6305–6316. doi:10.1021/bi970503q.
  4. 1 2 Kim, Jungwook; Raushel, Frank M. (2004). "Perforation of the Tunnel Wall in Carbamoyl Phosphate Synthetase Derails the Passage of Ammonia between Sequential Active Sites†". Biochemistry. 43 (18): 5334–5340. doi:10.1021/bi049945+. ISSN 0006-2960.
  5. ↑ ALTON MEISTER. “MECHANISM AND REGULATION OF THE GLUTAMINE-DEPENDENT CARBAMYL PHOSPHATE SYNTHETASE OF ESCHERZCHZA COLI.” Department of Biochemistry, Cornell University Medical College, New York, New York 10021
  6. ↑ MICHAEL KOTH*, BINNUR EROGLU, et al. “Novel mechanism for carbamoyl-phosphate synthetase: A nucleotide switch for functionally equivalent domains.” Department of Biology, Northeastern University, Boston, MA 02115 http://www.pnas.org/cgi/reprint/94/23/12348.pdf?ck=nck
  7. ↑ Thoden, James B.; Miran, Sophie G.; Phillips, James C.; Howard, Andrew J.; Raushel, Frank M.; Holden, Hazel M. (June 1998). "Carbamoyl Phosphate Synthetase: Caught in the Act of Glutamine Hydrolysis". Biochemistry. 37 (25): 8825–8831. doi:10.1021/bi9807761.
  8. ↑ David Nelson and Michael Cox. “Principles of Biochemistry, fourth edition.” Pgs 666-669

External links

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