In light of increasing global protein prices and with the need to reduce environmental impact of contemporary systems of milk production, the current study was performed to evaluate the effect of feeding on ration with different crude protein CP levels, in Holstein dairy cows ration on feed intake, milk production, reproductive performance and some hormonal and biochemical parameters in blood serum and milk. Two hundred and four Holstein dairy cows in three groups, 103, 63 and 38 were fed diets containing 20% CP, 18.4% CP and 17.5% CP for Groups 1, 2 and 3, respectively. Amino acid methionine and rumen undegredable protein (RUP) increased by decreasing the amount of CP. The average feed consumption and average milk production /head/day was reported. Gonadotrophin, Prostaglandin, Gonadotrophin protocol “GPG” were applied to the studied groups and artificial insemination were applied and ultrasonography was used to detect pregnancy and different reproductive findings. Blood and milk samples were collected to estimate serum progesterone hormone and milk urea nitrogen (MUN). Milk from each group was collected for analysis of milk compositions. The obtained results showed non-significant difference in the average dry matter intake per head per day between the studied groups (25.65 ± 0.92 kg), (24.45 ± 0.60 kg) and (24.62 ± 0.49 kg) respectively, with significant increase (P < 0.05) in the average milk production /head/day in group 3 (35.34 ± 1.27 kg) compared to group 1 (31.63 ± 0.57 kg). The analysis of milk samples revealed no significant difference in its constituents except in case of fat % which showed significant increase (P < 0.05) in group 3, (3.94 ± 0.04 %) compared with group 2, (3.70 + 0.05) also milk samples showed significant decrease (P < 0.05) in the average milk urea nitrogen concentration in group 3 (13.09 ± 1.15 mg/dl) compared to group 1 (16.64 ± 0.79 mg/dl). The obtained results showed highly significant increase (P < 0.01) in conception rate in groups 3 and 2 (39.47%) and (41.27%) respectively compared with group 1(17.48%). Serum progesterone level 12 days post-insemination showed significant increase (P < 0.05) in group 3 (2.06 ± 0.25 ng/ml) compared to group 1 (0.74 ± 0.11 ng/ml). This study concluded that reducing the CP level as in group 3 with adjusting the requirements of amino acids, energy and RUP improved fertility and profitability in Holstein dairy cows.