Protein Metabolism II ANS 520

Protein Pathways

Fate of Rumen Ammonia 1. Bacterial protein synthesis 2. Absorbed from reticulorumen and omasum NH 3 passes from rumen by diffusion into portal blood. (High concentration to low) Form of ammonia dependent on pH of rumen NH 3 + H + NH 4 + Less absorption at more acid pH 3. At pH of rumen, no NH 3 lost as gas

Fate of Absorbed Ammonia 1. Transported to liver by portal vein 2. Converted to urea via urea cycle in liver NH 3 Urea Urea cycle 3. Urea released into blood 4. If capacity of urea cycle in liver is exceeded Ammonia toxicity Over consumption of urea

Fate of Blood Urea 1. Excreted into urine 2. Recycled to digestive tract, g N/d • Saliva – Related to concentration of urea in blood Sheep: 0.5 to 1.0

Cattle: 1.0 to 7.6

• Diffusion into GIT Sheep: 2 to 5 Cattle: 25 to 40

Blood urea Urea Diffusion into Rumen Rumen wall Urea NH 3 1.

2.

Total N transferred is greater when high N diets are fed.

Percentage of diet N transferred is greater when low N diet are fed Bacterial population

Blood urea Urea Diffusion into Rumen Update Rumen wall Urea transporter Urea High [NH 3 ] inhibits NH 3 Bacterial population

Adjustments to Low Protein Intake Blood urea Kidney Urea Urine urea Urea is predominant form of N in urine Reabsorption of urea by kidney increased when ruminants fed low N diets • Conserves nitrogen in the body • Greater portion recycled to digestive tract • Sheep fed the same diet tend to reabsorb more urea than cattle

Nitrogen Recycling - Cattle

45 40 35 30 25 20 15 10 5 0 87.6

110.4

147.5

N intake, g/d 178.7

Marini et al. JAS 2003

203.5

GIT Saliva Wall

Sources of Nitrogen Recycled to GIT 1. Urea flowing back into digestive tract  Rumen • Saliva • Diffusion from blood  Lower digestive tract (large intestine, colon, • • cecum) Diffusion from blood Endogenous protein secretions into GIT    Mucins Enzymes Sloughing of tissue 2. Turnover of microbial cells in rumen & reticulum

Significance of Recycled Nitrogen Source of N for microbes when protein consumption is limited • Wild species Protein intake during winter is very low Rumen deficient of nitrogen for microbial activity • Slowly degraded feed proteins Recycling provides nitrogen for microbial growth • Infrequent feeding of supplemental protein • Programs to reduce supplemental nitrogen Difficult to make ruminants severely protein deficient

Urea Nitrogen - Cattle

Plasm urea Saliva urea 14 12 10 8 6 4 2 0 1.45

2.5

N, % Diet DM 3.4

Marini et al. JAS 2003

Urine N 140 120 100 80 60 40 20 0 Urine, Urea N N, % Diet DM

Amino Acid Synthesis Ammonia Fixation 1. Glutamine synthetase/glutamate synthase • Glutamine synthetase Glu + NH 3 + ATP Gln • Glutmate synthase  -ketoglutarate + glutamine + NADPH 2 2 Glu High affinity for NH 3 cells – Uses ATP - Concentrates NH 3 in Because of N recycling this reaction may not be that important

Amino Acid Synthesis Ammonia Fixation 2. Glutamic dehydrogenase •  -ketoglutarate + NH 3 + NADH Glu Low affinity for NH 3 – High concentration of enzyme in rumen bacteria – Does not use ATP Probably predominant pathway 3. Other AA can be synthesized by transamination reactions with glutamic acid Estimates of NH 3 requirements range from 5 (culture) to 20 mg/100 ml (in situ digestion)

Amino Acid Composition % Crude Protein or G/100g CP Methionine Lysine Histidine Phenylalanine Tryptophan Threonine Leucine Isoleucine Valine Arginine 1.97

6.37

2.47

3.53

0.49

3.90

6.70

2.84

4.03

3.30

Tissue Milk 2.71

7.62

2.74

4.75

1.51

3.72

9.18

5.79

5.89

3.40

----------Bact --------- Cell wall Non wall Mean 2.40

5.60

2.68

8.20

2.60

7.90

1.74

4.20

NA 3.30

5.90

4.00

4.70

3.82

2.69

5.16

1.63

5.59

7.51

5.88

6.16

6.96

2.00

5.10

5.80

8.10

5.70

6.20

5.10

Corn Soy 2.28

3.03

3.16

5.32

0.89

3.67

1.46

4.18

12.66 7.95

3.67

5.44

5.32

5.06

1.46

6.32

2.72

5.65

5.65

7.53

Amino Acids in Undegraded Feed Proteins Fish meal Fish meal residue His 3.4

2.9

Isl 4.2

4.9

Lys 6.6

6.0

Met 3.1

2.9

Sources of Amino Acids for Host Animal 1. Microbial proteins Quantity determined by: a) Fermentability of the feed b) Quantity of feed consumed c) Nitrogen available to microorganisms 2. Undegraded feed proteins (UIP) Quantity will vary in relation to: a) Degradability of feed proteins b) Quantity of feed proteins consumed

Nutritional Value of Microbial Proteins 1996 NRC for Beef Microbial protein 80% digestible in the intestine UIP 80% digestible in the intestine 2001 NRC for Dairy and Level 1 CNCPS Microbial protein 80% digestible in the intestine Digestibility of RUP (UIP) is variable in Dairy NRC UIP 80% digestible in Level 1 CNCPS

History of Protein Systems for Ruminants • ISU Metabolizable protein system • Wisconsin system – When urea could be used • Several European systems – Mostly MP systems • 1985 NRC system – Summarized systems & Proposed a MP system Used in 1989 Dairy NRC • Cornell CNCPS • 1996 Beef NRC system – Mostly CNCPS system Used in ISU Brands system • 2001 Dairy NRC system

Metabolizable Protein Model Tissue proteins A NH 3 B C Protein from diet NH Blood urea 3 Urine Amino acid pools Energy Microbial protein Protein Metabolizable protein Rumen Intestine Feces

Protein Metabolism of Ruminants Concept of Metabolizable Protein Metabolizable protein (MP) = Absorbed amino acids or = Digestible fraction of microbial proteins + digestible fraction of undegraded feed proteins Digestible protein (amino acids) available for metabolism Concept is similar to Metabolizable energy

Protein Metabolism in the Rumen

Less Extensively Degraded Protein

Feed Rumen Intestine Microbes Digestion Undegraded feed Metabolizable protein

Protein Metabolism in the Rumen

Extensively Degraded Protein

Feed Rumen Intestine Microbes NH

3

Digestion Undegraded feed Metabolizable protein

Metabolizable Protein Supply to Host Animal Metabolizable protein (MP): Microorganisms – Digestible proteins Undegraded feed proteins – Digestible proteins Microorganisms g/d = 0.13 (TDN intake, g/d) (0.8) (0.8) Microbes 80% true protein that is 80% digested Feed g/d = (Feed protein) (Portion undegraded) (0.8) Feed proteins 80% digested

Absorption of Amino Acids Amino acids and small peptides absorbed by active transport (specific for groups of AA) From intestines Portal blood Transport of amino acids into cells is similar process From blood Cells Active transport, requires energy

Utilization of Absorbed Amino Acids Via portal vein to liver • Used for synthesis of proteins in liver • Metabolized (deaminated) - Used for energy – Carbon for glucose • Escape the liver Carried by blood to body tissues • Used for synthesis of tissue proteins, milk, fetal growth, wool • Metabolized - Used for energy

Requirements for Absorbed Amino Acids Metabolizable Protein (MP) Protein (amino acid) requirements 1. Maintenance 2. Growth 3. Lactation 4. Pregnancy 5. Wool

Protein Metabolism Concept of Net Protein Net protein = protein gained in tissues, milk, or fetal growth = NP Metabolizable protein is used with less than 100% efficiency Net protein = (MP - Metabolic loss) As a quantity, net protein is less than metabolizable protein

Net Protein Required for Production Amino Acids Proteins Milk kg/d = (Milk yield, kg/d) (% protein in milk) Growth g/d = SWG (268 - (29.4 (RE/SWG))) SWG = Shrunk weight gain, kg/d RE = Retained energy, Mcal/d RE obtained from net energy equations .

Protein Metabolism Metabolic Loss Protein synthesis and metabolism of amino acids draw from the same pool Proteins Amino acids Metabolism • Metabolic loss results from continuous catabolism from amino acid pools • Continuous turnover of tissue proteins adds to amino acid pools in tissues

Amino Acid (MP) Requirements

Maintenance (three fractions) Protein required to support zero gain or production 1. Metabolism Milk Wool (Synthesis) Scurf Pregnancy Tissue proteins = Endogenous urinary N Metabolized Urine Amino acids Feces GIT (Degradation) 2. Proteins lost from body surface (hair, skin, secretions) = Scurf proteins 3. Proteins lost from undigested digestive secretions and fecal bacteria = Metabolic fecal N

Papers for Lab 4/8/10

• doi:10.2527/jas.2009-2218 – “Effects of partial ruminal defaunation on urea nitrogen recycling, nitrogen metabolism, and microbial nitrogen supply in growing lambs fed low or high dietary crude protein concentrations” • doi: 10.2527/jas.2005-614 – “Effects of ractopamine and protein source on growth performance and carcass characteristics of feedlot heifers”

Figure Assignments • Urea-N recycling paper

– Table 1-group discussion – Table 2-Jessica A.

– Table 3-Kenny B.

– Table 4-JJ G.

– Figure 1 and 2-Dan K.

– Table 5-Kim M.

– Table 6-Amir N.

Figure Assignments • Ractopamine paper

– Table 1-Jose N.

– Table 2-Danielle P.

– Table 3-Erin R.

– Table 4-Nathan U.