02 protein digestion (1)
DESCRIPTION
biochemistryTRANSCRIPT
Protein Digestion
Monogastric Protein Digestion Whole proteins are not absorbed
Too large to pass through cell membranes intact
Digestive enzymes Hydrolyze peptide bonds
Secreted as inactive pre-enzymes Prevents self-digestion
H3N+ C
HC
R
O
NH
CH
CO
RNH
CH
C
R
O
O–
Monogastric Protein Digestion Initiated in stomach
HCl from parietal cells Stomach pH 1.6 to 3.2 Denatures 40, 30, and 20 structures
Pepsinogen from chief cells
Cleaves at phenylalanine, tyrosine, tryptophan
Protein leaves stomach as mix of insoluble protein, soluble protein, peptides and amino acids
Aromatic amino acids
Pepsinogen
HClPepsin
Protein Digestion – Small Intestine
Pancreatic enzymes secreted Trypsinogen Chymotrypsinogen Procarboxypeptidase Proelastase Collagenase
Zymogens
Monogastric Digestion – Small Intestine
Zymogens must be converted to active form Trypsinogen Trypsin
Endopeptidase Cleaves on carbonyl side of Lys & Arg
Chymotrypsinogen Chymotrypsin Endopeptidase
Cleaves carboxy terminal Phe, Tyr and Trp
Procarboxypeptidase Carboxypeptidase
Exopeptidase Removes carboxy terminal residues
Enteropeptidase/Trypsin
Trypsin
Trypsin
Protein Digestion Small intestine (brush border)
Aminopeptidases Cleave at N-terminal AA
Dipeptidases Cleave dipeptides
Enterokinase (or enteropeptidase) Trypsinogen trypsin Trypsin then activates all the other enzymes
Trypsin Inhibitors Small proteins or peptides Present in plants, organs, and
fluids Soybeans, peas, beans, wheat Pancreas, colostrum
Block digestion of specific proteins Inactivated by heat
Protein Digestion Proteins are broken down to
Tripeptides Dipeptides Free amino acids
Free Amino Acid Absorption
Free amino acids Carrier systems
Neutral AA Basic AA Acidic AA Imino acids
Entrance of some AA is via active transport
Requires energy
Na+ Na+
Amino Acid Transporters – Brush Border Membrane
Transport system
Energy required
Substrates carried
LBIMINOy+
Bo,+
bo,+
NoYesYesNoYesNo
Leu, other neutralPhe, Tyr, Trp, Ile, Leu,
ValPro, Gly
Basic amino acidsMost neutral and basicMost neutral and basic
Peptide Absorption
Form in which the majority of protein is absorbed
More rapid than absorption of free amino acids
Active transport Energy required
Metabolized into free amino acids in enterocyte
Only free amino acids absorbed into blood
Absorption of Intact Proteins Newborns
First 24 hours after birth Immunoglobulins
Passive immunity Adults
Paracellular routes Tight junctions between cells
Intracellular routes Endocytosis Pinocytosis
Of little nutritional significance... Affects health (allergies and passive immunity)
In the Enterocytes…
First cells that can use the amino acids Transport into portal
blood Protein synthesis
Digestive enzymes Structure and growth
Energy
Stoll et al. (1998)
%
Groff & Gropper, 2000
*Whole proteins are nutritionally insignificant...
Basolateral Membrane Transport of
free amino acids only* Peptides are
hydrolyzed within the enterocyte
Transport mainly by diffusion and Na-independent carriers
Protein Transport in the Blood
Amino acids diffuse across the basolateral membrane Enterocytes portal blood liver
tissues Transported mostly as free amino acids
Liver Breakdown of amino acids Synthesis of non-essential amino acids
Groff & Gropper, 2000
Overview of Protein Digestion and Absorption in Monogastrics
Ruminant Protein Digestion
Ruminants can exist with limited dietary protein sources due to microbial protein synthesis Essential amino acids synthesized
Microbial protein is not sufficient during: Rapid growth High production
Protein in the Ruminant Diet Types of protein:
Dietary protein – contains amino acids Rumen Degradable Protein (RDP) – available for
use by rumen microbes Rumen Undegradable Protein (RUP) – escapes
rumen fermentation; enters small intestine unaltered
Varies with diet, feed processing Dietary non-protein nitrogen (NPN) – not
true protein; provides a source of nitrogen for microbial protein synthesis
Relatively CHEAP - decreases cost of protein supplementation
Ruminant Protein Feeding Feed the rumen microbes first (RDP)
Two counteractive processes in rumen Degradation of (dietary) protein Synthesis of microbial protein
Feed proteins that will escape fermentation to meet remainder of animal’s protein requirements
Escape protein, bypass protein, or rumen undegradable protein (RUP)
Aldehydes increase inter-protein cross-linking Heat treatment
Utilization depends on Digestibility of RUP source in the small intestine Protein quality
Protein Degradation in RumenFeedstuff % Degraded
in 2 hours
Urea 100
Alfalfa (fresh) 90
Wheat Grain 78
Soybean Meal 65
Corn Grain 48
Blood Meal 18
Rumen Protein Utilization Factors affecting ruminal degradation
Rate of passage Rate of passage degradation
Solubility in water Must be solubilized prior to degradation
Heat treatment Degradation
N (and S) availability Energy availability (carbohydrates)
Protein Fractions Dietary proteins classified based on
solubility in the rumen A
NPN, instantly solubilized/degraded B1 B2 B3
Potentially degradable C
Insoluble, recovered in ADF, undegradable
Ruminant Protein Digestion
Rumen microbes use dietary protein Creates difference between protein quality in
feed and protein actually absorbed by host Microbes break down dietary protein to
Amino acids NH3, VFAs, and CO2
Microbes re-synthesize amino acids Including all the essential amino acids from NH3 and
carbon skeletons
No absorption of protein or amino acids from rumen (or from cecum or large intestine!)
Protein Hydrolysis by Rumen Microbes Process with multiple steps
Insoluble protein is solubilized when possible Peptide bonds of solubilized protein are cleaved
Microbial endo- and exo-peptidases Amino acids and peptides released
Peptides and amino acids absorbed rapidly by bacteria
Bacteria degrade into ammonia N (NH3) NH3 used to produce microbial crude protein (MCP)
Microbial Crude Protein (MCP) Protein produced by microbial
synthesis in the rumen Primary source of protein to the
ruminant animal Microbes combine ammonia nitrogen
and carbohydrate carbon skeleton to make microbial crude protein
Diet affects the amount of nitrogen entering the small intestine as microbial crude protein
Factors Limiting Microbial Protein Synthesis Amount of energy
ATP Available nitrogen
NPN Degraded feed intake protein nitrogen (RDP)
Available carbohydrates Carbon residues for backbone of new amino acid
Microbial crude protein synthesis relies on synchronization of carbohydrate (for carbon backbones) and nitrogen availability (for amino group)
Microbial Protein Synthesis Synchronization of carbohydrate and N availability
NPN supplementation Carbohydrates used for carbon skeleton of amino acids
VFA (CHO fermentation)
Rumen NH3
Blood NH3
Adapted from Van Soest, 1994
Time post-feeding
Con
cent
ratio
n
Carbon backbone(from CHO fermentation)
Microbial Protein Formation
Dietary NPN
Dietary Soluble RDP
Microbial ProteinsAmino
Acids
Carbon Skeletons
Sulfur
Other Co-factors
NH3 ATP
Dietary Starch Sugar
Dietary Cellulose Hemicellulose
rapid
slow
rapid
slower
Dietary Insoluble RDP
very slow
Nitrogen Recycling Excess NH3 is absorbed
through the rumen wall to the blood Quickly converted to urea in the liver
Excess NH3 may elevate blood pH Ammonia toxicity Costs energy Urea (two ammonia molecules linked together)
Relatively non-toxic Excreted in urine Returned to rumen via saliva (rumination important)
Efficiency of nitrogen recycling decreases with increasing nitrogen intake
Nitrogen Recycling Nitrogen is continually recycled to
rumen for reutilization Ability to survive on low nitrogen diets Up to 90% of plasma urea CAN be recycled
to rumen on low protein diet Over 75% of plasma urea will be excreted
on high protein diet Plasma urea enters rumen
Saliva Diffuses through rumen wall from blood
Urea
Ammonia + CO2
Urease
Feed Protein, NPN and CHO
Feed Protein
Feed NPN
NH3/NH4
Bacterial N
NH4+ loss
MCP
RDP
RUPFeed Protein
AA
MCP
AA
NH3
Liver
Blood Urea
Salivary N
ATP
RUMEN
SMALL INTESTINE
Ruminant Digestion and Absorption
Post-ruminal digestion and absorption closely resembles the processes of monogastric animals However, amino acid profile entering
small intestine different from dietary profile
Overview of Protein Feeding Issues in Ruminants
Rumen degradable protein (RDP) Low protein quality in feed very good
quality microbial proteins Great protein quality in feed very good
quality microbial proteins Feed the cheapest RDP source that is
practical regardless of quality Rumen undegradable protein (RUP)
Not modified in rumen, so should be higher quality protein as fed to animal
May cost more initially, but may be worth cost if performance boosted enough
Salivary Urea
NPN
NH3
POOL
Dietary Nitrogen Non-utili
zed Ammonia
NH3 UREA
LIVER
LEVEL TOPROVIDE FORMAXIMUMMICROBIAL GROWTH
MICROBIAL PROTEIN
65% OF PROTEIN
35% OF PROTEIN
SMALL INTESTINE
AMINO ACIDS
AMINO ACIDSPROTEIN
AMINO ACIDS
PEPTIDES
Reticulo-rumen
RUP
RDP
Recycled urea
Functional Feeds
Functional feeds may be defined as any feed or feed ingredient that produces a biological effect or health benefit that is above and beyond the nutritive value of that feedstuff
Many feeds and their components fit this definition
Functional Proteins
Functional proteins are feed-derived proteins that, in addition to their nutritional value, produce a biological effect in the body
Feedstuffs with Biologically Active Proteins Milk Colostrum Whey Protein Concentrates/Isolates Plasma or serum Other animal-derived feedstuffs
Fish meal Meat and bone meal
Fermented animal-based products Yeast Lactobacillus organisms
Soy products
Protein Size Affects Function Many protein hormones are functional even
when fed to animals thyrotropin-releasing hormone (TRH, a 3-amino acid
peptide) luteinizing hormone-releasing hormone (LHRH, a 10-
amino acid peptide) insulin (a 51-amino acid polypeptide)
The smaller the peptide, the more “functional” it is when fed
100% activity for TRH, 50% for LHRH, and 30% for insulin Feedstuffs containing protein hormones
(colostrum) have biological activity when fed to animals
Production of Bioactive Peptides From Biologically-Inactive Proteins Peptides produced from intact inactive
proteins by incomplete digestion via proteases in stomach and duodenum or via microbial proteases in rumen
Many of these biologically active peptides (typically 2-4 amino acid residues) are stable from further digestion Some peptides bind to specific epithelial
receptors in intestinal lumen and induce physiological reactions
Some peptides are absorbed intact by a specific peptide transporter system into the circulatory system and transported to target organs
Responses to Feeding Functional Proteins or Peptides
Antimicrobial – including control of gut microflora Antiviral Binding of enterotoxins Anti-carcinogenic Immunomodulation Anti-oxidant effects Opioid effects Enhance tissue development or function Anti-inflammatory Appetite regulation Anti-hypertensive Anti-thrombic
Functional Activity of Major Milk Proteins Caseins (α, β and κ)
Transport of minerals and trace elements (Ca, PO4, Fe, Zn, Cu), precursor of bioactive peptides, immunomodulation (hydrolysates/peptides)
β-Lactoglobulin Retinol carrier, binding fatty acids, potential antioxidant, precursor for
bioactive peptides α-Lactalbumin
Lactose synthesis in mammary gland, Ca carrier, immunomodulation, anticarcinogenic, precursor for bioactive peptides
Immunoglobulins Specific immune protection (antibodies and complement system), G, M, A
potential precursor for bioactive peptides Glycomacropeptide
Antiviral, antithrombotic, bifidogenic, gastric regulation Lactoferrin
Antimicrobial, antioxidative, anticarcinogenic, anti-inflammatory, immunomodulation, iron transport, cell growth regulation, precursor for bioactive peptides
Lactoperoxidase Antimicrobial, synergistic effect with Igs and LF
Lysozyme Antimicrobial, synergistic effect with Igs and LF
Serum albumin Precursor for bioactive peptides
Proteose peptones Potential mineral carrier
Functional Activity of Minor Milk Proteins
Growth factors (IgF, TGF, EGF) stimulation of cell proliferation and differentation
Cytokines regulation of immune system (interferons,
interleukins, TGFβ, TNFα) Inflammation Increases immune response
Milk basic protein (MBP) Promotion of bone formation and suppression of
bone resorption Osteopontin
Modulation of trophoblastic cell migration
Protein Fragments That Have Biological Activity
Functional Protein Effects During Toxin or Disease Challenge
During intestinal inflammation, some functional proteins:
Reduce local inflammatory response excessive activation of inflammatory cells permeability
Increase Nutrient absorption Barrier function Intestinal health
During intestinal inflammation, some functional proteins:
Are absorbed and create adverse allergenic and immune responses in the body
Modified from Campbell, 2007