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Phenylketonuria (PKU) Phenylalanine hydroxylase is an enzyme which converts phenylalanine to tyrosine. A deficiency of this enzyme leads to a buildup of phenylalanine which results in severe mental retardation. Phenylalanine accumulates and is metabolized by alternate degradative pathway into phenylpyruvic acid and others leading to mental retardation.

Tyrosinemia There are three types of tyrosinemia. Each has distinctive symptoms and is caused by the deficiency of a different enzyme. Type I tyrosinemia, the most severe form of this disorder, is caused by a shortage of the enzyme fumarylacetoacetate hydrolase. Type II tyrosinemia is caused by a deficiency of the enzyme tyrosine aminotransferase. Diagnostic criteria include an elevated tyrosine level using MS/MS coupled with a confirmatory test for an elevated level of the abnormal metabolite succinylacetone

Cystinuria It is caused by a defect in the amino acid transport system rather than a metabolic enzyme deficiency. Normally, amino acids are free filtered by the glomerulus and then actively reabsorbed in the proximal renal tubules. In cystinuria, there is a 20-30 fold increase in the urinary excretion of cystine due to a genetic defect in the renal resorptive mechanism. Because cystine is relatively insoluble, it tends to precipitate in the kidney tubules and form urinary calculi. Cystinuria can be tested by cyanide-nitroprusside test. Ion exchange chromatography can be used for quantitative analysis of amino acids in urine or plasma.

Enzyme classification Plasma vs. non-plasma specific enzymes Plasma specific enzymes have a very definite/specific function in the plasma 1) Plasma is normal site of action 2) Concentration in plasma is greater than in most tissues 3) Often are liver synthesized 4) Examples: cholinesterase, plasmin, thrombin cholinesterase is an enzyme that catalyzes the hydrolysis of the neurotransmitter acetylcholine into choline and acetic acid, a reaction necessary to allow a cholinergic neuron to return to its resting state after activation. M. Zaharna Clin. Chem. 2015

Enzyme classification Non-plasma specific enzymes have no known physiological function in the plasma 1) Some are secreted into the plasma 2) A number of enzymes associated with cell metabolism normally found in the plasma only in low concentrations. An increased plasma concentration of these enzymes is associated with cell disruption or death M. Zaharna Clin. Chem. 2015

Factors Affecting Enzyme Levels in Blood Entry of enzymes into the blood Leakage from cells Altered production of enzymes E.g. increased osteoblastic activity results in increase in enzymes in bone disease Clearance of enzymes Half life vary from few hours to several days M. Zaharna Clin. Chem. 2015

Measuring enzyme activity Enzyme activity can be tested by measuring Increase of product Decrease of substrate Decrease of co-enzyme Increase of altered co-enzyme If substrate and co-enzyme are in excess concentration, the reaction rate is controlled by the enzyme activity. M. Zaharna Clin. Chem. 2015

Haptoglobin (α2) Synthesized in the liver Considered an acute-phase protein Binds free hemoglobin Prevents loss of Hemoglobin & Iron into urine Used to detect and evaluate hemolytic anemia and to distinguish it from anemia due to other causes Haptoglobin Reticulocytes 1 Decreased Increased Hemolytic anemia 2 Normal RBC destruction may be occurring in organs such as the spleen and liver 3 Anemia present is not due to RBC breakdown When the haptoglobin and hemoglobin attach, the reticuloendothelial cells (mainly in the spleen) remove the haptoglobin–hemoglobin complex from circulation 2- Because the freed hemoglobin is not released into the bloodstream, the haptoglobin is not used up and so is normal.

Ceruloplasmin (α2) Synthesized in the liver acute-phase reactant Copper carrying protein 90% of serum copper is bound to it Ordered along with blood and/or urine copper tests to help diagnose Wilson's disease, decreased levels of ceruloplasmin and excess storage of copper in the liver, brain, and other organs resulting in hepatic cirrhosis and neurologic damage. Wilson's disease, an autosomal recessive inherited disorder

α2 Macroglobulin Tetramer of four identical subunits Synthesized by liver major component of the α2 band in protein electrophoresis Primarily intravascular spaces due to size Inhibits proteases trypsin, pepsin & plasmin

Transferrin (Siderophilin) The major component of the -globulin. Glycoprotein synthesized by liver Carries 2 ferric iron molecules Normally 33% saturated Prevents loss of iron through kidneys Transports to storage sites where Iron is transferred to ferritin Transports to bone marrow for RBC synthesis Negative acute-phase protein

Hemopexin Beta globulin acute-phase reactant Purpose is to remove circulating Heme Breakdown product of Hemoglobin & myoglobin Carried to liver – broken down Increased in pregnancy, Some malignancies Low hemopexin levels are diagnostic of a hemolytic anemia Hemopexin can be determined by radial immunodiffusion

Disease Correlations Azotemia: elevated conc. of urea in blood Very high plasma urea concentration accompanied by renal failure is called uremia, or the uremic syndrome Causes of urea plasma elevations are: Prerenal Renal and postrenal

Pre-Renal Azotemia Reduced renal blood flow Less blood is delivered to the kidney less urea filtered Anything that produces a decrease in functional blood volume, include: Congestive heart failure, shock, hemorrhage, dehydration High protein diet or increased catabolism (Fever, major illness, stress)

Renal Azotemia Decreased renal function causes increased blood urea due to poor excretion Acute & Chronic renal failure Glomerular nephritis Tubular necrosis caused by a lack of oxygen to the kidney tissues (ischemia of the kidneys). & other Intrinsic renal disease Glomerulonephritis, also known as glomerular nephritis, abbreviated GN, is a renal disease characterized by inflammation of the glomeruli, or small blood vessels in the kidneys

Post-Renal Azotemia Obstruction of urine flow Renal calculi Tumors of bladder or prostate Severe infections

Uric Acid Uric acid is a final breakdown product of purine metabolism (adenosine/guanine) in liver Most other mammals degrade it further to allantoin Uric acid is transported to kidney and filtered (Renal excretion accounts for about 70% of uric acid elimination) 98% reabsorbed in proximal convoluted tubule (PCT) Some secreted by distal convoluted tubule (DCT) Net amount 6-12% of filtered amount Remaining 30% by GIT PCT= proximal convoluted tubule DCT= distal

Uric Acid Present in plasma as monosodium urate At plasma pH → relatively insoluble Conc. > 6.8 mg/dl → plasma saturated → urate crystals may form & precipitate in tissue Uric acid is measured to: assess inherited disorders of purine metabolism, to confirm diagnosis and monitor treatment of gout, to assist in the diagnosis of renal calculi, to prevent uric acid nephropathy during chemotherapeutic treatment, and to detect kidney dysfunction