Principles of Cheese Technology 241recombinant chymosin is a product of
fermentation technology, making it eco-
nomically reliable and acceptable to most
vegetarian, Jewish, Islamic, and Hindu con-
sumers. It replicates the quality of cheese
obtained by traditional calf rennet. Commer-
cial rennet preparations typically are 50%
calf chymosin and 50% bovine pepsin.
Calf rennet preparations are 95% chymosin.
Recombinant rennet preparations provide
more consistency in heat tolerance and a
dependable ratio of clotting activity and
general proteolytic activity.
The clotting of milk by rennin at normal
pH 6.6 is a three - phase reaction. In the
primary stage, rennet cleaves a specifi c bond
(phenyl alanine 105 - methionine 106 ) in the κ -
casein molecule, slicing it into para - κ - casein
and soluble glycomacropeptide fractions.
The hydrolyzed κ - casein can no longer hold
the hydrophobic casein particles together.
The Ca^ +^2 ions commence coagulation of
casein micelles in cheese milk when about
80% of the phenylalanine 105 - methionine 106
bonds are cleaved. In the secondary stage, the
micelles aggregate to form clusters, which
lead to gel formation. Water, along with
soluble constituents and fat, are trapped in
the three - dimensional network. In the fi nal
stage the network continues to attain fi rm-
ness. Cutting the gel is timed according to the
type of cheese being produced. In soft,
ripened cheeses, the gel is allowed to acquire
more fi rmness, while for hard cheeses, the
cutting process starts as soon as adequate
fi rmness is achieved.
Residual rennet in cheese curd plays an
important role in ripening. It is estimated that
less than 15% of rennet used in cheese
making is recovered in some varieties of
cheese curd. Calf rennet is destroyed by the
high cooking temperatures used in Swiss and
Italian cheeses, but in cheddar cheese a sig-
nifi cant amount of rennet survives and par-
ticipates in proteolysis to yield desirable
texture and fl avor. Microbial rennet prepara-
tions are usually more heat resistant and tendmode of coagulation in the vast majority
of the world ’ s cheeses. Normally, a casein
micelle is composed of calcium - sensitive
casein fractions ( α (^) s1 - , α (^) s2 - , and β - casein) pro-
tected by an envelope of calcium - insensitive
κ - casein. κ - casein is hydrolyzed as a result
of the enzyme action. Accordingly, calcium -
sensitive casein fractions are liberated and by
interacting with calcium of milk, they aggre-
gate and form coagulum.
As compared to other raw materials in
cheese manufacture, the quantity of coagulat-
ing enzymes is rather small. For most cheese
varieties, single - strength rennet has a usage
level of 200 ml/1,000 kg of milk and the
coagulation time is 20 to 30 minutes. The
coagulating enzymes ’ preparations are
obtained from animal, microbial, and vegeta-
ble sources, and they may take the form of a
crude paste, powder, or liquid rennet. Most
rennet preparations contain the coagulating
enzyme chymosin or rennin with varying
levels of pepsin.
Traditionally, rennet extracted from the
abomasum (fourth stomach) of milk - suckling
young calves was preferred. Because of scar-
city of the calf rennet, or to accommodate
religious beliefs, rennet - substitutes from
other sources have been developed, including
pepsins from swine, cows, and chickens.
Microbial sources include Mucor miehi,
Mucor pusillus , and Endothia parasitica.
Clotting activity is ascribed to specifi c action
on the phenyl alanine 105 - methionine 106 link-
age in the κ - casein molecule, whereas prote-
oltic activity refers to more random action.
Rennet substitutes generally display
excessive proteolytic activity, resulting in
reduced cheese yield and a bitter fl avor in
ripened cheeses. Synthetic chymosin is
obtained by recombinant DNA technology.
The genetic material from the calf abomas-
sum is transferred to a host organism —
Escherichi coli, Kluyvermices lactis, or
Aspergillus niger — where it is used as a
template to produce a coagulating enzyme
identical to calf chymosin. Therefore, the