In mammalian cells, the enzyme is responsible for converting (2R)-methylacyl-CoA esters to their (2S)-methylacyl-CoA epimers and known substrates, including coenzyme A esters of pristanic acid (mostly derived from phytanic acid, a 3-methyl branched-chain fatty acid that is abundant in the diet) and bile acids derived from cholesterol. This transformation is required in order to degrade (2R)-methylacyl-CoA esters by β-oxidation, which process requires the (2S)-epimer. The enzyme is known to be localised in peroxisomes and mitochondria, both of which are known to β-oxidize 2-methylacyl-CoA esters.
This enzyme belongs to the family of isomerases, specifically the racemases and epimerases which act on other compounds. The systematic name of this enzyme class is 2-methylacyl-CoA 2-epimerase. In vitro experiments with the human enzyme AMACR 1A show that both (2S)- and (2R)-methyldecanoyl-CoA esters are substrates and are converted by the enzyme with very similar efficiency. Prolonged incubation of either substrate with the enzyme establishes an equilibrium with both substrates or products present in a near 1:1 ratio. The mechanism of the enzyme requires removal of the α-proton of the 2-methylacyl-CoA to form a deprotonated intermediate (which is probably the enol or enolate) followed by non-sterespecific reprotonation. Thus either epimer is converted into a near 1:1 mixture of both isomers upon full conversion of the substrate.
Reduction of the protein level or activity results in the accumulation of (2R)-methyl fatty acids such as bile acids which causes neurological symptoms. The symptoms are similar to those of adult Refsum disease and usually appear in the late teens or early twenties.
The first documented case of AMACR deficiency was reported in 2006. This deficiency falls within a class of disorders called peroxisome biogenesis disorders (PBDs), although it is quite different from other peroxisomal disorders and does not share classic Refsum disorder symptoms. The deficiency causes an accumulation of pristanic acid, DHCA and EHCA and to a lesser extent Very long chain fatty acid and phytanic acid. This phenomenon was verified in 2002, when researchers reported of a certain case, "His condition would have been missed if they hadn't measured the pristanic acid concentration." 
AMACR deficiency can cause mental impairment, confusion, learning difficulties, and liver damage. It can be treated by dietary elimination of pristanic and phytanic acid through reduced intake of dairy products and meats such as beef, lamb, and chicken. Compliance to the diet is low, however, because of eating habits and loss of weight.
Increased levels of AMACR protein concentration and activity are associated with prostate cancer, and the enzyme is used widely as a biomarker (known in cancer literature as P504S) in biopsy tissues. Around 10 different variants of human AMACR have been identified from prostate cancer tissues, which variants arise from alternative mRNA splicing. Some of these splice variants lack catalytic residues in the active site or have changes in the C-terminus, which is required for dimerisation. Increased levels of AMACR are also associated with some breast, colon, and other cancers, but it is unclear exactly what the role of AMACR is in these cancers.
The enzyme is also involved in a chiral inversion pathway which converts ibuprofen, a member of the 2-arylpropionic acid (2-APA) non-steroidal anti-inflammatory drug family (NSAIDs), from the R-enantiomer to the S-enantiomer. The pathway is uni-directional because only R-ibuprofen can be converted into ibuprofenoyl-CoA, which is then epimerized by AMACR. Conversion of S-ibuprofenoyl-CoA to S-ibuprofen is assumed to be performed by one of the many human acyl-CoA thioesterase enzymes (ACOTs). The reaction is of pharmacological importance because ibuprofen is typically used as a racemic mixture, and the drug is converted to the S-isomer upon uptake, which inhibits the activity of the cyclo-oxygenase enzymes and induces an anti-inflammatory effect. Human AMACR 1A has been demonstrated to epimerise other 2-APA-CoA esters, suggesting a common chiral inversion pathway for this class of drugs.
^Sharma S, Bhaumik P, Schmitz W, Venkatesan R, Hiltunen JK, Conzelmann E, Juffer AH, Wierenga RK (Mar 2012). "The enolization chemistry of a thioester-dependent racemase: the 1.4 Å crystal structure of a reaction intermediate complex characterized by detailed QM/MM calculations". The Journal of Physical Chemistry. B116 (11): 3619–29. doi:10.1021/jp210185m. PMID22360758.
^Darley DJ, Butler DS, Prideaux SJ, Thornton TW, Wilson AD, Woodman TJ, Threadgill MD, Lloyd MD (Feb 2009). "Synthesis and use of isotope-labelled substrates for a mechanistic study on human alpha-methylacyl-CoA racemase 1A (AMACR; P504S)". Organic & Biomolecular Chemistry7 (3): 543–52. doi:10.1039/b815396e. PMID19156321.
^Ferdinandusse S, Denis S, Clayton PT, Graham A, Rees JE, Allen JT, McLean BN, Brown AY, Vreken P, Waterham HR, Wanders RJ (Feb 2000). "Mutations in the gene encoding peroxisomal alpha-methylacyl-CoA racemase cause adult-onset sensory motor neuropathy". Nature Genetics24 (2): 188–91. doi:10.1038/72861. PMID10655068.
^Rubin MA, Bismar TA, Andrén O, Mucci L, Kim R, Shen R, Ghosh D, Wei JT, Chinnaiyan AM, Adami HO, Kantoff PW, Johansson JE (Jun 2005). "Decreased alpha-methylacyl CoA racemase expression in localized prostate cancer is associated with an increased rate of biochemical recurrence and cancer-specific death". Cancer Epidemiology, Biomarkers & Prevention14 (6): 1424–32. doi:10.1158/1055-9965.EPI-04-0801. PMID15941951.
^>Zhou M, Jiang Z, Epstein JI (2003). "Expression and diagnostic utility of alpha-methylacyl-CoA-racemase (P504S) in foamy gland and pseudohyperplastic prostate cancer". Am. J. Surg. Pathol.27 (6): 772–8. doi:10.1097/00000478-200306000-00007. PMID12766580.
^Woodman TJ, Wood PJ, Thompson AS, Hutchings TJ, Steel GR, Jiao P, Threadgill MD, Lloyd MD (Jul 2011). "Chiral inversion of 2-arylpropionyl-CoA esters by human α-methylacyl-CoA racemase 1A (P504S)--a potential mechanism for the anti-cancer effects of ibuprofen". Chemical Communications47 (26): 7332–4. doi:10.1039/c1cc10763a. PMID21614403.
Jiang Z, Woda BA, Wu CL, Yang XJ (Aug 2004). "Discovery and clinical application of a novel prostate cancer marker: alpha-methylacyl CoA racemase (P504S)". American Journal of Clinical Pathology122 (2): 275–89. doi:10.1309/EJUY-UQPE-X1MG-68MK. PMID15323145.
Bautch S (Aug 1991). "Wisconsin doctor selected as national symbol of physicians' sacrifices". Wisconsin Medical Journal90 (8): 485–7. PMID1926890.
Schmitz W, Albers C, Fingerhut R, Conzelmann E (Aug 1995). "Purification and characterization of an alpha-methylacyl-CoA racemase from human liver". European Journal of Biochemistry / FEBS231 (3): 815–22. doi:10.1111/j.1432-1033.1995.tb20766.x. PMID7649182.
Maruyama K, Sugano S (Jan 1994). "Oligo-capping: a simple method to replace the cap structure of eukaryotic mRNAs with oligoribonucleotides". Gene138 (1-2): 171–4. doi:10.1016/0378-1119(94)90802-8. PMID8125298.
Suzuki Y, Yoshitomo-Nakagawa K, Maruyama K, Suyama A, Sugano S (Oct 1997). "Construction and characterization of a full length-enriched and a 5'-end-enriched cDNA library". Gene200 (1-2): 149–56. doi:10.1016/S0378-1119(97)00411-3. PMID9373149.
Ferdinandusse S, Denis S, Clayton PT, Graham A, Rees JE, Allen JT, McLean BN, Brown AY, Vreken P, Waterham HR, Wanders RJ (Feb 2000). "Mutations in the gene encoding peroxisomal alpha-methylacyl-CoA racemase cause adult-onset sensory motor neuropathy". Nature Genetics24 (2): 188–91. doi:10.1038/72861. PMID10655068.
Kotti TJ, Savolainen K, Helander HM, Yagi A, Novikov DK, Kalkkinen N, Conzelmann E, Hiltunen JK, Schmitz W (Jul 2000). "In mouse alpha -methylacyl-CoA racemase, the same gene product is simultaneously located in mitochondria and peroxisomes". The Journal of Biological Chemistry275 (27): 20887–95. doi:10.1074/jbc.M002067200. PMID10770938.
Amery L, Fransen M, De Nys K, Mannaerts GP, Van Veldhoven PP (Nov 2000). "Mitochondrial and peroxisomal targeting of 2-methylacyl-CoA racemase in humans". Journal of Lipid Research41 (11): 1752–9. PMID11060344.
Rubin MA, Zhou M, Dhanasekaran SM, Varambally S, Barrette TR, Sanda MG, Pienta KJ, Ghosh D, Chinnaiyan AM (Apr 2002). "alpha-Methylacyl coenzyme A racemase as a tissue biomarker for prostate cancer". JAMA287 (13): 1662–70. doi:10.1001/jama.287.13.1662. PMID11926890.
Luo J, Zha S, Gage WR, Dunn TA, Hicks JL, Bennett CJ, Ewing CM, Platz EA, Ferdinandusse S, Wanders RJ, Trent JM, Isaacs WB, De Marzo AM (Apr 2002). "Alpha-methylacyl-CoA racemase: a new molecular marker for prostate cancer". Cancer Research62 (8): 2220–6. PMID11956072.
Zhou M, Chinnaiyan AM, Kleer CG, Lucas PC, Rubin MA (Jul 2002). "Alpha-Methylacyl-CoA racemase: a novel tumor marker over-expressed in several human cancers and their precursor lesions". The American Journal of Surgical Pathology26 (7): 926–31. doi:10.1097/00000478-200207000-00012. PMID12131161.
Varambally S, Dhanasekaran SM, Zhou M, Barrette TR, Kumar-Sinha C, Sanda MG, Ghosh D, Pienta KJ, Sewalt RG, Otte AP, Rubin MA, Chinnaiyan AM (Oct 2002). "The polycomb group protein EZH2 is involved in progression of prostate cancer". Nature419 (6907): 624–9. doi:10.1038/nature01075. PMID12374981.
Leav I, McNeal JE, Ho SM, Jiang Z (Mar 2003). "Alpha-methylacyl-CoA racemase (P504S) expression in evolving carcinomas within benign prostatic hyperplasia and in cancers of the transition zone". Human Pathology34 (3): 228–33. doi:10.1053/hupa.2003.42. PMID12673556.
Shen-Ong GL, Feng Y, Troyer DA (Jun 2003). "Expression profiling identifies a novel alpha-methylacyl-CoA racemase exon with fumarate hydratase homology". Cancer Research63 (12): 3296–301. PMID12810662.
Zha S, Ferdinandusse S, Denis S, Wanders RJ, Ewing CM, Luo J, De Marzo AM, Isaacs WB (Nov 2003). "Alpha-methylacyl-CoA racemase as an androgen-independent growth modifier in prostate cancer". Cancer Research63 (21): 7365–76. PMID14612535.